Biological Fluid Filtration System

ABSTRACT

A biological fluid filtration system for scanning a biological fluid so as to filter potentially undesirable constituents from the biological fluid for therapeutic or diagnostic purposes. The biological fluid filtration system generally includes a fluid receiving device adapted to receive a biological fluid. A valve including an inlet, a first outlet, and a second outlet is fluidly connected to the fluid receiving device. The biological fluid within the fluid receiving device is scanned by a scanner to produce scanned data relating to the biological fluid. A control unit in communication with the scanner and the valve receives the scanned data and controls the valve based on the scanned data. The valve is controlled to direct the biological fluid through either the first or second outlet of the valve depending upon the constituents of the biological fluid identified by the control unit.

CROSS REFERENCE TO RELATED APPLICATIONS

The present application is a continuation of U.S. application Ser. No.17/162,450 filed on Jan. 29, 2021 (Docket No. PART-004), which claimspriority to U.S. Provisional Application No. 62/968,476 filed Jan. 31,2020 (Docket No. PART-002). Each of the aforementioned patentapplications, and any applications related thereto, is hereinincorporated by reference in their entirety.

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT

Not applicable to this application.

BACKGROUND Field

Example embodiments in general relate to a biological fluid filtrationsystem for optically scanning a biological fluid so as to filterpotentially harmful constituents from the biological fluid fortherapeutic or diagnostic purposes.

Related Art

Any discussion of the related art throughout the specification should inno way be considered as an admission that such related art is widelyknown or forms part of common general knowledge in the field.

Eight million people die from cancer each year and it is predicted that13.2 million patients will do so in 2030. 90% of deaths are caused bymetastases. Cancer metastasis spreads when Circulating Tumor Cells(CTCs) dislodge from the primary tumor site and travel via thecirculatory system (or lymphatic system) and lodge at another site.Certain cancer types disseminate as single cells, while others—such asoral squamous cell carcinoma, colorectal carcinoma, melanoma, breastcancer, endometrial carcinoma, and pancreatic cancer, do so bycollective cell (CTC-cluster) migration. Significant variability hasalso been found from transcriptional profiling and surface markeranalysis, leading to an understanding that CTCs are highlyheterogeneous. In 2012, it was shown that CTC-variability is highlyconsistent with tumor tissue variability, meaning that phenotyping ofCTCs will bring to light many of the characteristics present in thetumor itself.

It is estimated that every day, tumors release thousands of cells intothe circulation where CTCs survive for about 1-2.5 hours. Most CTCsundergo apoptosis upon release or remain dormant, and only a few (0.1%)are able to survive the effect of stressors and ultimately then formdistant metastases. Using the only FDA-approved system for CTCseparation, CellSearch (Menarini Silicon Biosystems), researchersdetected a single CTC (median 1, range from 0 to 4) in 7.5 mL of bloodin 13% of analyzed samples, while analyzing 30-mL blood volume samplesallowed a detection rate of 2 CTCs (range from 0 to 9) in 47% of theanalyzed samples. Thus, rarity is a significant challenge for theidentification and separation of CTCs and CTC-clusters. 10 mL of aperipheral blood sample from a metastatic cancer patient typicallycontains 0-100 single CTCs and roughly 0-5 CTC-clusters (5-20% of allCTCs) among approximately 50×10⁹ red blood cells, 80×10⁶ white bloodcells and 3×10⁹ platelets. Furthermore, some CTCs overlap in size withwhite blood cells, making size based separation challenging.

Yet, due to the clinical significance of CTCs in cancer prognosis andtreatment, research in techniques to separate CTCs is very active.Separation strategies are categorized as positive enrichment, negativeenrichment, and label-free techniques. Positive enrichment typicallyrefers to a process that selects for the CTCs while leaving the otherparticles behind. This typically has a very high accuracy, and one ofthe most common methods involves antibody tagging surface antigens (e.g.EpCAM) on the CTC.

Negative enrichment involves targeting and removing other types ofcells, in this case, white and red blood cells. This generally leads tolower purity but does not have the challenge of removal of bindingprobes from the surface of the CTC, and is often able to bypass thechallenge of CTC heterogeneity. A label-free technique is one thatavoids biochemically tagging the desired molecule. This means thatrather than using immune affinity for capture or sample cleaning,another method that does not involve labeling cells is used such assize-based, mechanical property based, acoustic, or optical. Someresearchers have proposed combining several techniques, such as opticaldetection of fluorescently tagged CTCs followed by size basedmicrofluidic separation, to achieve better results. Recently, buildingupon advances in AI and machine learning, methods have been published toautomatically detect CTCs by training machine learning algorithms onmicroscopic images of CTCs in sample blood.

SUMMARY

An example embodiment is directed to a biological fluid filtrationsystem. The biological fluid filtration system generally includes afluid receiving device adapted to receive a biological fluid. A valveincluding an inlet, a first outlet, and a second outlet is fluidlyconnected to the fluid receiving device. The biological fluid within thefluid receiving device is scanned by a scanner to produce scanned datarelating to the biological fluid. A control unit in communication withthe scanner and the valve receives the scanned data and controls thevalve based on the scanned data. The valve is controlled to direct thebiological fluid through either the first or second outlet of the valvedepending upon the constituents of the biological fluid identified bythe control unit.

There has thus been outlined, rather broadly, some of the embodiments ofthe biological fluid filtration system in order that the detaileddescription thereof may be better understood, and in order that thepresent contribution to the art may be better appreciated. There areadditional embodiments of the biological fluid filtration system thatwill be described hereinafter and that will form the subject matter ofthe claims appended hereto. In this respect, before explaining at leastone embodiment of the biological fluid filtration system in detail, itis to be understood that the biological fluid filtration system is notlimited in its application to the details of construction or to thearrangements of the components set forth in the following description orillustrated in the drawings. The biological fluid filtration system iscapable of other embodiments and of being practiced and carried out invarious ways. Also, it is to be understood that the phraseology andterminology employed herein are for the purpose of the description andshould not be regarded as limiting.

BRIEF DESCRIPTION OF THE DRAWINGS

Example embodiments will become more fully understood from the detaileddescription given herein below and the accompanying drawings, whereinlike elements are represented by like reference characters, which aregiven by way of illustration only and thus are not limitative of theexample embodiments herein.

FIG. 1 is a perspective view of a biological fluid filtration system inaccordance with an example embodiment.

FIG. 2 is a perspective view of a biological fluid filtration system inaccordance with an example embodiment.

FIG. 3 is a perspective view of a biological fluid filtration system inaccordance with an example embodiment.

FIG. 4 is a perspective view of a biological fluid filtration system inaccordance with an example embodiment.

FIG. 5 is a perspective drawing of a fluid receiving device withmicrofluidic channels in accordance with an example embodiment.

FIG. 6 is a schematic drawing of a fluid receiving device withmicrofluidic channels in accordance with an example embodiment.

FIG. 7 is a schematic drawing of a fluid receiving device withmicrofluidic channels in accordance with an example embodiment.

FIG. 8 is a perspective drawing of a fluid receiving device with amicrowell array in accordance with an example embodiment.

FIG. 9 is a schematic drawing of a fluid receiving device with amicrowell array in accordance with an example embodiment.

FIG. 10 is a schematic drawing of a fluid receiving device with amicrowell array in accordance with an example embodiment.

FIG. 11 is a histogram of typical cell count data in blood.

FIG. 12 is a block diagram of an aphaeretic system and method ofremoving undesirable constituents using microfluidic channels inaccordance with an example embodiment.

FIG. 13 is a block diagram of an aphaeretic system and method ofremoving undesirable constituents using microwell arrays in accordancewith an example embodiment.

FIG. 14 is a block diagram of a diagnostic system in accordance with anexample embodiment.

FIG. 15 is a block diagram of an aphaeretic system and method ofremoving undesirable pathogens from component blood fluid in accordancewith an example embodiment.

FIG. 16 is a block diagram of an aphaeretic system and methodincorporating image data of known pathogens and optical artifacts into areference data in accordance with an example embodiment.

FIG. 17 is a block diagram of an aphaeretic system and method ofremoving undesirable CTCs from blood in accordance with an exampleembodiment.

FIG. 18 is a block diagram of an aphaeretic system and methodincorporating image data of CTCs and CTC-clusters into a reference datain accordance with an example embodiment.

FIG. 19 is a block diagram of an aphaeretic system and method ofremoving undesirable CTCs from whole blood fluid in accordance with anexample embodiment.

FIG. 20 is a block diagram of an aphaeretic system and method ofincorporating image data of CTCs and CTC-clusters into a reference datain accordance with an example embodiment.

FIG. 21 is a block diagram of an aphaeretic system and method forremoval of undesirable CTCs and CTC-clusters using blood samples fromrepresentative individuals other than the patient to obtain a referencedata in accordance with an example embodiment.

FIG. 22 is a block diagram of an aphaeretic system and methodincorporating image data of pathogens into a reference data inaccordance with an example embodiment.

FIG. 23 is a block diagram of filtration of cytokines from bloodcomponent fluid in accordance with an example embodiment.

FIG. 24 is a block diagram of immunomagnetic filtration of CTCs fromblood component fluid in accordance with an example embodiment.

FIG. 25 is a block diagram of molecular and genetic profiling ofundesirable CTCs removed from blood in accordance with an exampleembodiment.

FIG. 26 is a graphical display of cell characteristics based on imagedata of cells.

FIG. 27 is a block diagram of an aphaeretic system and method ofcomputer processing of reference image data obtained from optic scans ofbiological fluid components from fluid samples of representativeindividuals other than patient.

FIG. 28 is a block diagram of an aphaeretic system and method ofcomputer processing of reference image data obtained from optic scans ofbiological fluid components from fluid sample of patient.

FIG. 29 is a block diagram of an aphaeretic system and method whichutilizes a certifying/verification algorithm to remove undesirablepathogens from component blood fluid in accordance with an exampleembodiment.

FIG. 30 is a flowchart illustrating a system and method of filtering abiological fluid utilizing both a pre-filtration session and afiltration session in accordance with an example embodiment.

FIG. 31 is a block chart illustrating a scanner utilizing digitalholographic microscopy of a biological fluid filtration system inaccordance with an example embodiment.

FIG. 32 is a flowchart illustrating a system and method of filtering abiological fluid for use in connection with leukapheresis in accordancewith an example embodiment.

FIG. 33 is a flowchart illustrating a system and method of filtering abiological fluid for use in connection with the removal of stem cells inaccordance with an example embodiment.

FIG. 34 is a block diagram illustrating a system and method of filteringa biological fluid for use in connection with testing of drugs andtreatments in accordance with an example embodiment.

FIG. 35 is a flowchart illustrating a multi-stage system and method offiltering a biological fluid in accordance with an example embodiment.

FIG. 36A is a block diagram illustrating a system and method offiltering a biological fluid including a fluid receiving deviceutilizing droplet sorting in accordance with an example embodiment.

FIG. 36B is a block diagram illustrating additional channels added afteran isolation path of a biological fluid filtration system in accordancewith an example embodiment.

FIG. 37 is a flowchart illustrating a system and method of filtering abiological fluid for use in connection with filtration of subsets ofhealthy cells in accordance with an example embodiment.

FIG. 38 is a flowchart illustrating a system and method of filtering abiological fluid utilizing a presorting stage in accordance with anexample embodiment.

FIG. 39 is a flowchart illustrating a system and method of filtering abiological fluid utilizing a presorting stage and having multiple outletports in accordance with an example embodiment.

FIG. 40 is a flowchart illustrating a system and method of filtering abiological fluid utilizing a presorting stage for diagnostics inaccordance with an example embodiment.

FIG. 41 is a flowchart illustrating a system and method of filtering abiological fluid utilizing a presorting stage for diagnostics withhealthy cells being returned in accordance with an example embodiment.

FIG. 42 is a flowchart illustrating a system and method of filtering abiological fluid utilizing a presorting stage for diagnostics withhealthy cells being returned in accordance with an example embodiment.

FIG. 43 is a block diagram illustrating a system and method of filteringa biological fluid utilizing multiple pre-sorting stages in accordancewith an example embodiment.

FIG. 44A is a block diagram illustrating a body-worn device of abiological fluid filtration system worn on a patient in accordance withan example embodiment.

FIG. 44B is a block diagram illustrating a body-worn device of abiological fluid filtration system including anti-coagulant and bufferfluid worn on a patient in accordance with an example embodiment.

FIG. 44C is a perspective view of a portable filtration device of abiological fluid filtration system in accordance with an exampleembodiment.

FIG. 45 is a block diagram illustrating a body-worn device of abiological fluid filtration system in accordance with an exampleembodiment.

FIG. 46 is a flowchart illustrating the usage of a body-worn device of abiological fluid filtration system in accordance with an exampleembodiment.

FIG. 47 is a flowchart illustrating the usage of a body-worn device witha pre-sorting stage of a biological fluid filtration system inaccordance with an example embodiment.

FIG. 48 is a flowchart illustrating the usage of a body-worn device withdrug infuser in the return path of a biological fluid filtration systemin accordance with an example embodiment.

FIG. 49 is a block diagram illustrating a closed-loop filtering,monitoring, and testing system of a biological fluid filtration systemin accordance with an example embodiment.

FIG. 50 is a block diagram illustrating a closed-loop filtering,monitoring, and testing system of a biological fluid filtration systemutilizing drug choice optimization in accordance with an exampleembodiment.

FIG. 51 is a block diagram illustrating a biological fluid filtrationsystem being connected to a dialysis unit in accordance with an exampleembodiment.

FIG. 52 is a block diagram illustrating a biological fluid filtrationsystem being connected to a blood bag in accordance with an exampleembodiment.

FIG. 53 is a block diagram illustrating the usage of a body-worn deviceto filter and sequester small samples of blood containing CTCs andCTC-clusters of a biological fluid filtration system in accordance withan example embodiment.

DETAILED DESCRIPTION A. Overview

An example biological fluid filtration system 10 generally comprises areceiver path 20 adapted to receive a biological fluid 16 from abiological fluid source 17. The receiver path 20 may comprise varioustypes of conduits, channels, or the like known in the art fortransferring a fluid between two locations such as, for example, anapheresis catheter, an indwelling line, or a venous catheter. Otherexamples could include two intravenous (IV) lines: one for access to anartery and the other for access to a vein. The systems and methodsdescribed herein may be utilized for filtering a wide range ofbiological fluids 16, such as but not limited to blood, lymphatic fluid,cerebrospinal fluid, sweat, urine, pericardial fluid, stools, saliva,and the like. The biological fluid source 17 may comprise the patient 12herself or, in some embodiments, a separate reservoir in which thebiological fluid 16 is temporarily stored after being drawn from thepatient 12.

As shown throughout the figures, a fluid receiving device 30 may befluidly connected to the receiver path 20 so as to receive thebiological fluid 16 from the receiver path 20. The fluid receivingdevice 30 generally comprises a structure in which the biological fluid16 is temporarily stored or channeled to be optically scanned by ascanner 70. As discussed in more detail below, the fluid receivingdevice 30 may vary in different embodiments. In a first exemplaryembodiment, the fluid receiving device 30 may comprise one or moremicrofluidic channels 31. In a second exemplary embodiment, the fluidreceiving device 30 may comprise a microwell array 32 comprised of aplurality of microwells. In yet another exemplary embodiment, the fluidreceiving device 30 may comprise a microfluidic droplet generator 34 toscan cell encapsulating droplets 39.

A valve 40 may be positioned downstream of the fluid receiving device30, with the valve 40 being configured to direct flow of the biologicalfluid 16 after being scanned within the fluid receiving device 30. Thevalve 40 may comprise an inlet 41 which is fluidly connected to thefluid receiving device 30. The valve 40 may comprise a first outlet 42which is fluidly connected to an isolation path 50 and, in someembodiments, a second outlet 43 which is fluidly connected to a returnpath 60. In some embodiments, the return path 60 may be omitted, withthe biological fluid 16 being returned to the biological fluid source 17through the receiver path 20 rather than a separate return path 60. Insome embodiments, instead of or in addition to a return path 60 back tothe biological fluid source 17, a reprocessing path 62 may be utilizedto return back to the fluid receiving device 30 for further processing.

It should be appreciated that the valve 40 may have a default state inwhich flow is directed towards one of the two outlets 42, 43 in theabsence of control of the valve 40. For example, the valve 40 may be aspring-based valve 40 that, by default, directs flow to the first outlet42. Such a valve 40 may be adjusted to direct flow to the second outlet43, such as by activation of a spring. Upon release of the spring, sucha valve 40 may revert back to its original, default state in which thevalve 40 directs flow to the first outlet 42. Thus, the valve 40 may notneed to be controlled if the default state of the valve 40 alreadydirects flow to the desired outlet 42, 43.

The isolation path 50 is fluidly connected to the first outlet 42 of thevalve 40 such that the biological fluid 16 contents of the fluidreceiving device 30 may be isolated or sequestered from the biologicalfluid source 17 if undesirable constituents 14 are identified in thebiological fluid 16 during scanning by the scanner 70 as discussed inmore detail below. The isolation path 50 may comprise one or morechannels which are fluidly connected to the first outlet 42 of the valve40 such that any scanned biological fluid 16 containing undesirableconstituents 14 may be sequestered or isolated. The isolation path 50may be fluidly connected to a reservoir, cartridge, container, vessel,or any device capable of holding a fluid. Such sequestered or isolatedbiological fluids 16 may be utilized for diagnostics or may be disposedof.

The scanner 70 is generally oriented toward the fluid receiving device30. Although the singular term is used throughout, it should beappreciated that, in some embodiments, multiple scanners 70 may beutilized. For example, in an embodiment in which the fluid receivingdevice 30 is comprised of a microwell array 32, a first scanner 70 couldbe oriented toward a first portion of the microwell array 32 and asecond scanner 70 could be oriented toward a second portion of themicrowell array 32. In any case, the scanner 70 is adapted to opticallyscan the biological fluid 16 within the fluid receiving device 30 so asto derive a scanned data 90 of the biological fluid 16.

In an exemplary embodiment, the scanner 70 may utilize digitalholographic microscopy (DHM) to scan the contents of the fluid receivingdevice 30. In such an embodiment, the scanner 70 may comprise a lightsource 72 which is adapted to illuminate the biological fluid 16 to bescanned. More specifically, the scanner 70 may comprise a light source72 such as a laser or a light-emitting diode. In addition to the lightsource 72, the scanner 70 in such an embodiment will generally include amicroscope objective 73 adapted to gather light from the biologicalfluid 16 and an image sensor 76 so as to produce a holographic image.Thus, the microscope objective 73 will generally comprise a lens whichfunctions to collect an object wave 77 front created by the light source72 as discussed below.

The control unit 80 in such an embodiment may then function as a digitallens to calculate a viewable image of the object wave 77 front. Sincethe microscope objective 73 in such a digital holographic microscopyembodiment is only used to collect light, rather than to form an image,the microscope objective 73 may be omitted entirely in some embodiments.Other example embodiments could rely on other arrangements to generateholograms of the object being scanned.

A control unit 80 may be communicatively connected to the scanner 70 soas to receive the scanned data 90 of the biological fluid 16 from thescanner 70. The control unit 80 may be operatively connected to thevalve 40 such that the control unit 80 may direct the opening or closingof the inlet 41 and outlets 42, 43 of the valve 40.

The control unit 80 is adapted to compare the scanned data 90 of thebiological fluid 16 with a reference data 91. As discussed below, thereference data 91 may comprise patterns, criteria, images, and/or othercharacteristics of desirable constituents 15 such as red blood cells,white blood cells, and the like. The desirable constituents 15 maycomprise biological fluid constituents 13 that are desirable for aparticular patient 12 or for a particular application. It should beappreciated that a biological fluid constituent 13 which is desirablefor a first patient 12 may be undesirable for a second patient 12.Further, a biological fluid constituent 13 which is desirable for aparticular application may be undesirable for other applications.

Thus, the control unit 80 may be configured to determine if a particularsample of biological fluid 16 consists only of such desirableconstituents, with the sample of biological fluid 16 being directedthrough the first outlet 42 of the valve 40 if only desirableconstituents are identified. If the sample of biological fluid 16 doesnot consist exclusively of such desirable constituents, the sample ofthe biological fluid 16 may be directed instead through the secondoutlet 43 of the valve 40.

The control unit 80 is adapted to direct flow of the biological fluid 16from the fluid receiving device 30 by operation of the valve 40. If thescanned data 90 of the biological fluid 16 contains only desirableconstituents 15 exhibiting criteria that match with any of the healthyor desirable constituents of the reference data 91, the control unit 80will switch the valve 40 so as to direct the biological fluid 16 to thebiological fluid source 17 by a return path 60, such as by the receiverpath 20 in reverse. If the scanned data 90 of the biological fluid 16includes one or more undesirable constituents 14 not exhibiting criteriathat match with any known desirable constituents 15 of the referencedata 91, the control unit 80 will switch the valve 40 so as to directthe biological fluid 16 to the isolation path 50 to be isolated orsequestered from the biological fluid source 17 for further diagnosticor therapeutic processing. Alternatively, if the scanned data 90includes one or more undesirable constituents 14, the valve 40 may beswitched so as to direct the biological fluid 16 back to the fluidreceiving device 30 by a reprocessing path 62 for further processing.The scanned and reference data 90, 91 could include images and/ormorphological data of the cells in the images.

In an exemplary embodiment, the biological fluid filtration system 10may comprise a fluid receiving device 30 adapted to receive a biologicalfluid 16 from a biological fluid source 17. A valve 40 may be fluidlyconnected to the fluid receiving device 30, with the valve 40 includingan inlet 41, a first outlet 42, and a second outlet 43. The inlet 41 ofthe valve 40 is fluidly connected to the fluid receiving device 30. Ascanner 70 is configured to scan the biological fluid 16 within thefluid receiving device 30 to produce a scanned data 90 relating to thebiological fluid 16 within the fluid receiving device 30. A control unit80 is in communication with the scanner 70 and the valve 40, with thecontrol unit 80 being configured to receive the scanned data 90 from thescanner 70 and to control the valve 40 based on the scanned data 90 fromthe scanner 70. The control unit 80 is configured to control the valve40 to (a) direct the biological fluid 16 through the first outlet 42 ifthe control unit 80 determines that the scanned data 90 indicates apresence of an undesirable constituent 14 within the biological fluid 16or (b) direct the biological fluid 16 through the second outlet 43 ifthe control unit 80 determines that the scanned data 90 does notindicate a presence of an undesirable constituent 14 within thebiological fluid 16.

The fluid receiving device 30 may comprise a microfluidic channel 31. Aninlet valve 33 may be fluidly connected to an inlet of the microfluidicchannel 31 for pausing flow of the biological fluid 16 into themicrofluidic channel 31. The undesirable constituent 14 may be comprisedof an unknown constituent of the biological fluid 16 that is notrecognized by the control unit 80. The undesired constituent 14 may becomprised of a tumor cell. The control unit 80 may be configured tocompare the scanned data 90 to a reference data 91. The reference data91 may be comprised of a characteristic or an image of a healthy cell.The scanned data 90 may be comprised of an image.

The fluid receiving device 30 may comprise a plurality of microfluidicchannels 31. Each of the microfluidic channels 31 may be arranged inparallel. The scanner 70 may be configured to scan each of the pluralityof microfluidic channels 31. The biological fluid 16 may be comprised ofblood, cerebrospinal fluid, or lymphatic fluid. The first outlet 42 ofthe valve 40 may be fluidly connected to a reservoir. The second outlet43 of the valve 40 may be fluidly connected to a biological fluid source17. The scanner 70 may be comprised of an optical scanner such as adigital holographic microscope. The scanner 70 may include amonochromatic laser.

In an exemplary embodiment of the biological fluid filtration system 10,the control unit 80 is configured to control the valve 40 to (a) directthe biological fluid 16 through the first outlet 42 if the control unitdetermines that the scanned data 90 indicates that the biological fluid16 consists of only desirable constituents 15 or (b) direct thebiological fluid 16 through the second outlet 43 if the control unit 80determines that the scanned data 90 does not indicate that thebiological fluid 16 consists of only desirable constituents 15. Amicrofluidic separation module 100 may be fluidly connected to an inletof the fluid receiving device 30 for removing one or more biologicalfluid constituents 13 from the biological fluid 16 prior to scanning.

In an exemplary embodiment, the control unit 80 is configured todetermine that the scanned data 90 indicates the presence of theundesirable constituent 14 when the control unit 80 detects theundesirable constituent 14 within the scanned data 90. In anotherexemplary embodiment, the control unit 80 is configured to determinethat the scanned data 90 indicates the presence of the undesirableconstituent 14 when the control unit 80 detects an unknown constituentwithin the scanned data 90. In another exemplary embodiment, the controlunit 80 is configured to determine that the scanned data 90 indicatesthe presence of the undesirable constituent 14 when the control unit 80detects only desirable constituents 15 within the scanned data 90.

An exemplary method of filtering a biological fluid 16 using thebiological fluid filtration system 10 may comprise the steps ofreceiving the biological fluid 16 by the fluid receiving device 30;scanning the biological fluid 16 within the fluid receiving device 30 bythe scanner 70; directing the biological fluid 16 through the firstoutlet 42 if the control unit 80 determines that the scanned data 90indicates a presence of an undesirable constituent 14 within thebiological fluid 16; and directing the biological fluid 16 through thesecond outlet 43 if the control unit 80 determines that the scanned data90 indicates a presence of an undesirable constituent 14 within thebiological fluid 16.

B. Biological Fluid Filtration System

As shown in FIG. 30, an exemplary embodiment of a biological fluidfiltration system 10 may comprise an automated optofluidic system forremoval of undesirable constituents 14 from a biological fluid 16 byoptically inspecting the biological fluid constituents 13 andsequestering any undesirable constituents 14 that do not meet recognizedcriterial of known desirable constituents 15 of the biological fluid 16such as biological fluid constituents 13 that are desirable for aparticular patient 12 or application. It should be appreciated that thebiological fluid constituents 13 may comprise biological constituents ormay comprise non-biological constituents such as dyes and the like.

The systems and methods described herein may be utilized for thefiltration of a wide range of biological fluids 16, such as but notlimited to blood, lymphatic fluid, cerebrospinal fluid, sweat, urine,pericardial fluid, stools, saliva, and any other organism-derivedextracellular fluid that retains or transports nutrients, cells, wasteproducts, or foreign bodies, or that is susceptible to pathogenicinfection. It should be appreciated that the systems and methodsdescribed herein are further not intended to be limited to humans, andcould be utilized in connection with veterinary treatment of a widerange of animals in some embodiments.

The systems and methods described herein may be utilized for bothdiagnostic and therapeutic applications. In some embodiments, multiplebiological fluids 16 may be processed from the same individual (animalor human). As a non-limiting example, there are cases in which cancerhas spread from the original tumor site (e.g., breast, lung, etc.) tothe meninges surrounding the brain and/or spinal cord. In such cases,the systems and methods described herein may be utilized to process andfilter CTCs and CTC-clusters from multiple biological fluids 16, such asfrom blood as well as from cerebrospinal fluid. In some embodiments,these multiple biological fluids 16 may be aphaeretically processedsimultaneously, particularly in a diagnostic setting.

After sequestration of the output, the sequestered output may bereprocessed additional times using an optical filtration system (withoutthe aphaeretic components) to further isolate CTCs, CTC-clusters, WBC's,and cell-free plasma. By utilizing multiple valves and output ports, theisolated CTCs, CTC-clusters, WBC's, and cell-free plasma could be heldseparately for further processing.

In some embodiments, rather than the biological fluid 16 being directlysourced from a person or animal, the biological fluid 16 could be asample with or without a return path 60 back to its source. Suchembodiments could include the use in diagnostic systems to separatepathogens, CTCs, or CTC-clusters from biological samples for downstreamdiagnostics (e.g., genomic, transcriptomic, metabolomics, drugsensitivity, drug resistance, etc.) and characterization.

In another exemplary embodiment, the biological fluid source 17 could bean aphaeretic extract from a therapeutic aphaeresis or leukapheresismachine. Such leukapheresis machines are routinely utilized in labs,such as where WBC's are separated from a patient. In the case of cancerpatients, such leukapheresis extracts may contain CTCs or CTC-clustersand some platelets in addition to WBC's. FIG. 32 illustrates anexemplary method of optical filtration being utilized to processleukapheresis extracts and filter CTCs and CTC-clusters.

In yet another exemplary embodiment, tumor cell contaminants may beidentified and removed from autologous stem-cell transplant products.Autologous stem cell transplants are typically used in patients who needto undergo high doses of chemotherapy and radiation to cure theirdiseases. These treatments could be toxic and damage the bone marrow. Anautologous stem cell transplant aids to replace the damaged bone marrow,but it is often reported that the process to collect stem cells from thepatient could lead to contamination of such products with tumor cells.FIG. 33 illustrates an exemplary method of optical filtration beingutilized to process stem cell extracts.

Biological fluid constituents 13 of the biological fluid 16 may comprisedesirable constituents 15 and undesirable constituents 14. The desirableconstituents 15 may comprise entities within the biological fluid 16that are recognized as being normally found in the subject biologicalfluid 16 or that are desirable for a particular patient 12 orapplication. Such desirable constituents 15 will typically play a rolein the proper functioning of tissues, organs, and body systems. In someembodiments, desirable constituents 15 may comprise biological fluidconstituents 13 which are known to be benign or otherwise non-harmful.

The undesirable constituents 14 may comprise any biological fluidconstituents 13 which are not recognized as being benign, healthy, orotherwise non-harmful. Undesirable constituents 14 may comprise anyentity that is not a healthy constituent for a particular patient 12. Itshould be appreciated that biological fluid constituents 13 may behealthy or desirable for a first patient 12 and unhealthy or undesirablefor a second patient 12. Thus, the identification of a particularbiological fluid constituent 13 as either healthy/desirable orunhealthy/undesirable will typically be on a patient-by-patient basis.By way of example and without limitation, undesirable constituents 14may comprise circulating tumor cells (CTCs), CTC-clusters, andpathogens, including bacterial and fungal organisms, protozoa,extracellular vesicles, lipids, cholesterol, dyes, drugs, and infectiousviral agents. In some embodiments, any biological fluid constituent 13which is unrecognized or unknown may be assumed to be an undesirableconstituent 14.

FIG. 1 is a schematic diagram illustrating a configuration of oneembodiment of biological fluid filtration system 10. The illustratedexemplary embodiment includes a receiver path 20 adapted to receive abiological fluid 16 from a biological fluid source 17; a fluid receivingdevice 30 fluidly connected to the receiver path 20 so as to receive thebiological fluid 16 from receiver path 20; a valve 40 comprising aninlet 41, a first outlet 42, and a second outlet 43, wherein the inlet41 is fluidly connected to the fluid receiving device 30; an isolationpath 50 fluidly connected to first outlet 42; a return path 60 in fluidcommunication with second outlet 43; a scanner 70 oriented toward thefluid receiving device 30, and a control unit 80 communicativelyconnected to the scanner 70 and operatively connected to the valve 40.

The scanner 70 is adapted to optically scan the biological fluid 16within the fluid receiving device 30 so as to derive a scanned data 90of the biological fluid 16 and relay the scanned data 90 to the controlunit 80. The control unit 80 is adapted to compare the scanned data 90of the biological fluid 16 with a reference data 91, the reference data91 comprising recognized data patterns characteristic of desirableconstituents 15 of the biological fluid 16. In some embodiments, boththe scanned and reference data 90, 91 may comprise images which arecompared to each other. In other embodiments, the scanned and referencedata 90, 91 may comprise characteristics such as cellularcharacteristics including but not limited to cell solidity, cell surfacecharacteristics, nucleus-cytoplasmic ratio, convexity, luminescence,circularity, elongation, size, inner diameter and roundness. In otherembodiments, the scanned and reference data 90, 91 may comprise bothimages as well as characteristics of the cells in those images, such ascellular characteristics including but not limited to cell solidity,cell surface characteristics, nucleus-cytoplasmic ratio, convexity,luminescence, circularity, elongation, size, inner diameter androundness.

The control unit 80 is adapted to switch the valve 40 so as to directthe biological fluid 16 to the biological fluid source 17 by the returnpath 60 if the scanned data 90 of the biological fluid 16 includes onlydesirable constituents 15 having one or more recognized characteristicsof any of the desirable constituents 15 of the reference data 91. Thecontrol unit 80 is further adapted to switch the valve 40 so as todirect the biological fluid 16 to the isolation path 50 if the scanneddata 90 of the biological fluid 16 includes one or more undesirableconstituents 14 lacking one or more recognized characteristics of any ofthe desirable constituents 15 of the reference data 91. In someembodiments, a reprocessing path 62 may be included to route thebiological fluid 16 back to the fluid receiving device 30 for furtherprocessing. The return path 60 and reprocessing path 62 may comprise anydevice or conduit suitable for transferring a fluid.

The return path 60 may comprise a channel or a plurality of channelsthrough which the biological fluid 16 may be returned to the biologicalfluid source 17. For example, the return path 60 may comprise acatheter, one or more intravenous lines, an indwelling line, or a venouscatheter. Similarly, the reprocessing path 60 may comprise a channel ora plurality of channels through which the biological fluid 16 may bereturned to the fluid receiving device 30 for further processing.

The receiver path 20 includes any suitable vessel for sterile deliveryor transfer of a biological fluid 16 for various purposes such asclinical analysis or processing. For example, in the aphaeretic systemof the preferred embodiment, the receiver path 20 may be an apheresiscatheter, two intravenous lines (one for the artery and the other for avein), an indwelling line, or a venous catheter.

The fluid receiving device 30 includes any microfluidic device or systemsuitable for optic imaging or microscopy. The fluid receiving device 30will generally include an inlet through which biological fluid 16 isreceived into the fluid receiving device 30 and an outlet through whichbiological fluid 16 may exit the fluid receiving device 30. In someembodiments, the fluid receiving device 30 may include multiple inletsand/or multiple outlets. The inlet of the fluid receiving device 30 maybe fluidly connected to the biological fluid source 17, and the outletof the fluid receiving device 30 may be fluidly connected to anisolation path 50, return path 60, reprocessing path 62, or cartridge126, 146.

In one embodiment, which is shown schematically in FIGS. 5-7, the fluidreceiving device 30 comprises one or more microfluidic channels 31. Inthe illustrated embodiment, the microfluidic channels 31 are configuredfor illustration purposes as a batch of planar, parallel channels with alinear geometry. However, microfluidic channels suitable for use withsystems and methods described herein may have any number oftopographies, geometries and patterns. For example, the microfluidicchannels 31 may be etched or molded in a microfluidic chip.

In an alternative embodiment, which is shown in FIGS. 8 and 9, the fluidreceiving device 30 comprises a microwell array 32. Other suitablemicrofluidic devices include, e.g., glass capillary systems. In afurther embodiment, best shown in FIG. 36A, the fluid receiving device30 may comprise a microfluidic droplet generator 34.

The valve 40 is preferably a router-type micro-valve, but could includeother types as well such as diaphragm-type micro-valve withelectromagnetic actuation, piezo electric, thermoplastic, etc. The valve40 is actuated by a control signal 81 sent by the control unit 80. Thevalve 40 can be actuated by various methods such as, for example,mechanically, electrically, piezo, electro-thermally, pneumatically,electromagnetically, by phase changes, or by introduction of externalforce. The valve 40 is preferably a diaphragm-type micro-valve withelectromagnetic actuation. For fluid receiving devices 30 comprisingmultiple microfluidic channels 31 or a microwell array 32, acorresponding number of valves 40, in which each valve 40 exclusivelydirects the flow of contents out of a single corresponding channel orwell, is preferred. However, in some example embodiments, multiple wellscould share a valve 40. In other example embodiments, rather than valves40, pipettes could be utilized to extract contents from microwells.

The isolation path 50 includes any suitable vessel or microwells forsterile delivery or transfer of a biological fluid 16 for variouspurposes such as in connection with clinical analysis or processing. Ina preferred embodiment, biological fluid 16 directed to the isolationpath is transported and stored in accordance with appropriate hematologyprocedures for diagnostic or conformational analysis, as described inmore detail below.

The scanner 70 may comprise any device capable of scanning a biologicalfluid 16. The scanner 70 may comprise an optical scanner, including butnot limited to a digital holographic microscope. The scanner 70generally comprises a light source and optical detector. The scanner 70is preferably configured to develop a multi-modal reference data 91 fromwhich patterns characteristic of desirable constituents 15 of biologicalfluid 16 may be can be recognized. In an example embodiment, the scanner70 comprises a Differential Interference Contrast Microscopy System (DICMicroscopy System). Generally, DIC Microscopy generates contrast bytranslating refractive index gradients of different areas of a specimeninto amplitude variations that are visualized as differences inbrightness. In that regard, the DIC Microscopy System is adapted tocreate enhanced contrast images, and is particularly suited for use inimaging unstained cell specimens exhibiting little natural visiblecontrast. DIC Microscopy's contrast-enhanced imaging yields informationconcerning cell characteristics highly relevant for many of theembodiments presented herein, including but not limited to cellsolidity, cell surface characteristics, nucleus-cytoplasmic ratio,convexity, luminescence, circularity, elongation, size, inner diameterand roundness.

Other imaging devices and techniques suitable for use individually or incombination as a scanner 70, for example include, Phase contrastmicroscopy (PCM), Hoffman modulation, polarized light microscopy,holographic microscopy, confocal scanning optic microscopy (CSOM), orlaser scanning optic microscopy (SOM) to measure voxel fluorescence,bright-field microscopy, dark-field illumination, Raman spectrometry tomeasure Raman Scattering, Optical interferometry to measure opticalinterference, total internal reflection fluorescence microscopy tomeasure evanescent effect, planar waveguides for refractive indexdetection, photonic crystal biosensors for measure of biomolecules oncell surfaces, and light property modulation detections such as surfaceplasmon resonance (SPR) detection.

The optical detector of the scanner 70 may comprise a single lens, ormultiple linear lenses or lens arrays, and may include lenses ofdifferent shapes, for example, PCX lenses or Fresnel lenses.

In alternative embodiment, the scanner 70 and fluid receiving device 30are combined in individual modules in a modular optofluidic system(MOPS), which permits modification or reconfiguration of modules to suita given application.

The control unit 80 may be local, remote, or cloud-hosted such asthrough distributed networking. As shown in FIG. 29, the control unit 80follows an image processing software program comprising an algorithm 82(or series of algorithms) adapted to identify patterns in reference data91 of images taken of desirable constituents 15 of a biological fluid 16and to recognize those patterns in processing the scanned data 90obtained by the scanner 70.

If all biological fluid constituents 13 of the scanned biological fluid16 present in the fluid receiving device 30 at that moment sufficientlymatch one or more characteristics of desirable constituents 15recognized by the algorithm 82, then the image processing softwareprogram generates a first control signal instruction 83 to the controlunit 80 to relay a control signal 81 to the valve 40 to route thescanned biological fluid 16 to the return path 60.

If, on the other hand, one or more of the constituents 13 of the scannedbiological fluid 16 present in the fluid receiving device 30 at thatmoment fails to sufficiently match patterns of desirable constituents 15recognized by algorithm 82, then the image processing software programgenerates a second control signal instruction 84 to the control unit 80to relay a control signal 81 to the valve 40 to route the scannedbiological fluid 16 to the isolation path 50.

The control unit 80 may optionally also follow an additional imageprocessing software program adapted to recognize and search for patternsin the scanned data 90 indicating the presence of an undesirableconstituent 14 such as a disease-related constituent in the scannedbiological fluid 16. The additional processing software programpreferably comprises a certifying algorithm 85 adapted to providevalidation for each control signal instruction based on a verificationcondition. For example, a control signal instruction to route thescanned biological fluid 16 to the return path 60 is validated only oncondition that no patterns were detected in the scanned data 90 toindicate the presence of an undesirable constituent 14 such as atargeted disease-related constituent in the scanned biological fluid 16.If the condition is not satisfied, a corrected instruction is issued toroute the scanned biological fluid 16 to the isolation path 50.

In one embodiment, the reference data 91 is derived based on normativedata for recognized desirable constituents 15 of the biological fluid 16within a relevant reference population. In another embodiment, thereference data 91 is derived by imaging biological fluid 16 samplesobtained from representative individuals in the relevant referencepopulation. In other embodiments, the reference data 91 may at leastpartially be derived from a pre-filtration session with a particularpatient 12.

In such an embodiment, as illustrated in FIGS. 12-17 and 19, thereference data 91 is obtained based on imaging of a fluid sample of thesubject patient's biological fluid system during a pre-filtrationsession. Because the reference data 91 of the embodiment is patientspecific, data ambiguity based on constituent heterogeneity is avoided.Accordingly, the algorithm 82, which may rely upon machine learning, isconfigured to more precisely identify patterns of desirable constituents15 in the subject-specific reference data 91 and thus more preciselyrecognize desirable constituents 15 of the scanned data 90 taken of thesubject biological fluid source 17 during filtration.

In some embodiments, machine learning and/or artificial intelligencemodels may be utilized to develop and optimize the reference data 91. Byway of example, samples of biological fluids 16 from the patient 12 orfrom others may be analyzed to identify desirable constituents 15 of thebiological fluid 16 through machine learning. Such a system may be moreeasily accomplished with relation to healthy cells which are common toall patients 12, such as red blood cells, white blood cells, and thelike. This is a result of consistency of the characteristics or imagesof such healthy cells and the availability of a large sample size. Withrespect to undesirable constituents 14, machine learning and/orartificial intelligence models may also be applied, though with asmaller sample size and with the caveat that certain unhealthy cellssuch as CTCs may have different characteristics in different patients12. Additionally, the scanned data 90 may also be analyzed using machinelearning and/or artificial intelligence models. Further, the process ofcomparing the scanned data 90 with the reference data 91, and theresulting determination by the control unit 80, may also utilize machinelearning and/or artificial intelligence models.

In another embodiment, as illustrated in FIG. 16, the reference data 91includes imaging data allowing the control unit 80 to discern opticalartifacts, e.g., halos, air bubbles, or blurring, to avoid errors wherecharacteristics of such artifacts in the imaging data might otherwisetrigger an erroneous second control signal instruction 84 to directfluid containing only desirable constituents 15 to the isolation path50. For the same reason, the reference data 91 may include imaging dataallowing the control unit 80 to discern non-targeted biologicalentities, e.g., benign bacteria.

In another embodiment, the biological fluid filtration system 10conducts a continuous-flow microfluidic operation. In such anembodiment, the biological fluid 16 may continuously flow through thefluid receiving device 30 as the biological fluid 16 is scanned. In analternative embodiment, as illustrated in FIGS. 12-16, the microfluidicchannel 31 includes an inlet valve 33 such as a channel lock adapted tosuspend fluid flow while biological fluid 16 in the microfluidic channel31 is scanned.

In yet another embodiment, which is illustrated in FIGS. 2, 15, 17, 18,and 20-22, the biological fluid filtration system 10 performspre-processing on the biological fluid 16 using a microfluidicseparation module 100 to separate particular cellular constituents ofthe biological fluid 16 for promotion to the fluid receiving device 30.Suitable pre-processing devices and techniques suitable may rely on avariety of cell-characteristics to perform separation including, size,density, inertial hydrodynamic, antigen binding affinity, acoustics,motility, electric charge, electric dipole moment, centrifugation, ormagnetism. Some techniques might also involve the use of buffersolutions.

In one embodiment, in which blood is filtered for removal of CTCs, wholeblood is pre-processed to separate leukocytes from smaller-sized bloodconstituents (i.e., erythrocytes, thrombocytes, plasma) and large CTCsor CTC-clusters. The separated, leukocyte-enriched blood, which containssmall CTCs, is promoted to the fluid receiving device 30 for scanningand filtering. CTC-free leukocytes along with the other separatedhealthy blood constituents are returned to the subject's circulatorysystem, and the separated large and small CTCs or CTC-clusters aresequestered.

The other separated blood constituents can optionally be subjected toadditional filtering before being returned to the subject's circulatorysystem. For example, as illustrated in FIGS. 23-26, separated smallblood constituents can be subject to antigen-based filtration to removeproteins that promote metastasis, including aiding in CTC extravasationand evasion of immune response, e.g., cytokines, chemokines, and growthfactors released by tumor-associated macrophages. Antigen-basedfiltration may also be optionally used to compliment both pre-processingand optical scanning and filtering processes to remove any remainingCTCs in pre-processed or optically scanned and filtered biologicalfluids 16 before being returned to the subject.

In an exemplary embodiment, which is shown in FIGS. 1-4, the biologicalfluid 16 is blood, the biological fluid source 17 is the patient 12, andthe biological fluid filtration system 10 is an aphaeretic system thatcirculates blood from the circulatory system of a patient 12, removesundesirable constituents 14 from the blood, and then returns thefiltered, healthy blood back to the circulatory system. Such abiological fluid filtration system 10 has equal application to otherbiological fluids 16, including, for example, cerebrospinal fluid,lymphatic fluid, and the various other fluids described herein.

The biological fluid filtration system 10 can be applied as atherapeutic, a diagnostic system, or both. In one embodiment, apatient's pathology is determined and the particular cancer-associatedCTCs or disease causative agent identified, and the system is adapted tofilter out the known CTCs or agents from patients the biological fluidsystem. In such an embodiment as illustrated in FIGS. 3 and 4, thereceiver path 20 is adapted to receive and facilitate flow of biologicalfluid 16 from the patient 12 via, e.g., a cannula and venous catheterassembly, to the fluid receiving device 30.

As shown in FIGS. 1 and 2, the scanner 70 is adapted to optically scanthe biological fluid 16 on or within the fluid receiving device 30 so asto derive a scanned data 90 of the biological fluid 16 and relay thescanned data 90 to the control unit 80. The control unit 80 is adaptedto compare the scanned data 90 of the biological fluid 16 with areference data 91 to relay a control signal 81 to the valve 40 to routehealthy biological fluid 16 to the return path 60 and sequesterbiological fluid 16 containing CTCs or undesirable disease causativeagents to the isolation path 50, as described herein. The return path 60is adapted to receive and facilitate flow of healthy biological fluid 16from the fluid receiving device 30 via the valve 40, and returning thefluid 16 to the patient 12 via, e.g., a cannula and venous catheterassembly. In another embodiment, the sequestered biological fluid 16 isdiagnostically processed for conformational analysis.

In another embodiment, which is shown in FIG. 14, a subject's pathologyis undetermined, and the system is adapted to filter out undesirableconstituents 14 not recognized by the system as meeting pre-determinedcriteria characteristic of desirable constituents 15 of the subjectbiological fluid 16, as described herein. As with the therapeutic systemdescribed above, the scanner 70 of the diagnostic system is adapted tooptically scan the biological fluid 16 within the fluid receiving device30 so as to derive the scanned data 90 of the biological fluid and relaythe scanned data 90 to the control unit 80, and, in turn, the controlunit 80 is adapted to compare the scanned data 90 of the biologicalfluid 16 with the reference data 91 and relay a control signal 81 to thevalve 40 to route healthy biological fluid 16 to the return path 60 andsequester biological fluid 16 containing undesirable constituents 14,such as undesirable, indeterminate biological agents, as describedherein. If used for diagnostic purposes, in some embodiments, such asystem could be used for early detection of the signs of cancer in thepatient. In such diagnostic systems, even the filtered fluid containinghealthy components may not be returned back to the subject.

The sequestered biological fluid 16 is then subject to diagnostictesting to identify the undesirable constituents 14 present in thesequestered biological fluid 16. In the diagnostic embodiment, thereceiver path 20 is optionally adapted to receive and facilitate flow ofbiological fluid 16 from the patient's biological fluid system and thereturn path 60 is optionally adapted to return healthy biological fluid16 to the patient's biological fluid system. In one embodiment, thereceiver path 20 and return path 60 are respectively adapted to receiveand return biological fluid 16 from the subject's biological fluidsystem, the diagnostic system is adapted to both diagnose and filter outundesirable constituents 14 in the subject's biological fluid system. Inan alternative embodiment, the receiver path 20 is adapted to receiveand facilitate flow of a biological fluid 16 sample taken of thesubject's biological fluid system, and the healthy biological fluid 16routed to the return path 60 is processed for storage or disposal.

In some embodiments such as shown in FIG. 36B, the isolation path 50 mayfurther bifurcate into additional channels and valves 40, with each suchbifurcated additional channel including its own fluid receiving device30 and scanner 70. The geometry of such additional channels could be ofvarious shapes, including without limitation rectangular, serpentine,spiral, circular, square, or combinations thereof. The geometry of theexemplary embodiment shown in FIG. 36B should thus not be construed aslimiting.

Continuing to reference FIG. 36B, additional channels may be added afterthe isolation path 50 of any of the embodiments described and shownherein. Contents from such an isolation path 50 are scanned in multipleadditional stages for further enrichment of the biological fluid 16. Asshown in FIG. 36B, contents from an isolation path 50 enter anadditional fluid receiving device 30 and are scanned again by a scanner70. The resulting scanned data 90 is processed by the control unit 80.Valves 40 are provided to direct the contents into additional stages,with the valves 40 being operated by the control unit 80 based on theresults of its processing of the contents. In exemplary embodimentsutilized in an aphaeretic setting, any contents including only healthycells (e.g., desirable constituents 15) are carried forward, such asback to the biological fluid source 17 via a return path 60.

Contents which include a mixture of healthy and non-healthy cells (e.g.,any contents including undesirable constituents 14) are directed alongan additional channel to an additional fluid receiving device 30 and arescanned again by a scanner 70. After the additional fluid receivingdevice 30, valves 40 are provided to bifurcate the additional channelsuch that contents including only healthy cells may be diverted towardsa return path 60. Contents including non-healthy cells such asundesirable constituents 14 are directed along an additional channel toan additional fluid receiving device 30 and are scanned again by ascanner 70. This process may be repeated any number of times, withhealthy cells being carried forward to be isolated or returned, and anyunhealthy cells being scanned additional times.

As shown in FIG. 36B, pumps 46 may be utilized to regulate flow rates ofthe biological fluid 16 through the various channels. The pumps 46 maycomprise pressure pumps, syringes, peristaltic, and the like. Thescanning techniques could vary across successive stages or sections(e.g., DHM, digital inline holography, etc.) and the algorithms utilizedto identify cells may also vary (e.g., neural network type classifiersto decision trees, etc.). In other example embodiments, such anarrangement could involve droplets 39 as opposed to cells in media.

C. Fluid Receiving Device

As described and shown herein, a wide range of fluid receiving devices30 may be utilized for the biological fluid filtration system 10 incombination with various types of biological fluids 16, including butnot limited to blood and/or leukapheresis extracts. The fluid receivingdevice 30 may comprise a microfluidic platform. In a first exemplaryembodiment as shown in FIG. 5, the fluid receiving device 30 maycomprise one or more microfluidic channels 31. In a second exemplaryembodiment as shown in FIG. 8, the fluid receiving device 30 maycomprise a microwell array 32. In a third exemplary embodiment as shownin FIG. 36A, the fluid receiving device 30 may comprise a microfluidicdroplet generator 34. It should also be appreciated that various othertypes of fluid receiving devices 30 may be utilized.

It should be appreciated that, in some embodiments, multiple types offluid receiving devices 30 may be grouped together. For example, anexemplary fluid receiving device 30 may comprise both microfluidicchannels 31 and a microwell array 32, arranged together in parallel orin series. As a further example, a fluid receiving device 30 maycomprise both microfluidic channels 31 and a droplet generator 34. Asyet another example, a fluid receiving device 30 may comprisemicrofluidic channels 31, a microwell array 32, and a droplet generator34 all configured to work in concert. Additionally, in some embodiments,there could be multiple successive valves 40 and fluid receiving devices30 attached to a single outlet to enrich the contents of a microchannel.

As shown in FIG. 5, an exemplary embodiment of the fluid receivingdevice 30 may comprise one or more microfluidic channels 31. Themicrofluidic channels 31 will generally comprise networks of one ormultiple channels through which biological fluids 16 may be routed to beprocessed using the systems and methods described herein. In someembodiments, a single microfluidic channel 31 may be utilized, such aswith an embodiment effectuating single-stage processing of a singlebiological fluid 16. In other embodiments, multiple microfluidicchannels 31 may be utilized, such as with an embodiment effectuatingmulti-stage processing of a single biological fluid 16, or single-stageprocessing of multiple biological fluids 16 simultaneously.

By way of example, multiple fluid receiving devices 30 may be arrangedin parallel so as to allow for multiple biological fluids 16, eitherfrom the same or different biological fluid sources 17, to besimultaneously scanned and then routed accordingly. In this manner,multiple types of biological fluids 16 from the same or multiplebiological fluid sources 17 may be processed simultaneously. By way ofexample, a first biological fluid 16 may be processed in a first fluidreceiving device 30 and a second biological fluid may be processed in asecond fluid receiving device 30. Additional fluid receiving devices 30may similarly be operated in parallel to suit any number of biologicalfluids 16.

As a further example, multiple fluid receiving devices 30 may bearranged in series such that a single biological fluid 16, or multiplebiological fluids 16, may be scanned multiple times in multiple stages.For example, a first fluid receiving device 30 may process a biologicalfluid 16 a first time, and then the biological fluid 16 may betransferred to a second fluid receiving device 30 in series with thefirst fluid receiving device 30 so as to process the biological fluid 16a second time. Subsequent fluid receiving devices 30 may also be added,allowing for multi-stage separation of the biological fluid 16 asdiscussed herein. In some embodiments, multiple fluid receiving devices30 may be arranged both in parallel and in series so as to allowsimultaneous, multi-stage processing of multiple biological fluids 16.

The microfluidic channels 31 may be fabricated from polymeric materials,including but not limited to polymethylmethacrylate, cylic olefincopolymer (COC), polycarbonate, polydimethylsiloxane (PDMS), SU-8photoresist, and the like. As previously discussed, the microfluidicchannels 31 may comprise various topographies, geometries, and patterns,and thus should not be construed as limited by the exemplarytopographies, geometries, and patterns shown in the exemplary figures.

The microfluidic channels 31 may be arranged in a multilayer,2-dimensional or 3-dimensional configuration. Moreover, various channelgeometries may be employed including, for example, curvilinear andspiral geometries. Various channel patterns in addition to parallelpatterns may be suitable including, for example, knot, basket-weave, andbraided patterns. By way of example and without limitation, themicrofluidic channels 31 may be straight, curved, or helical. Wheremultiple microfluidic channels 31 are utilized, the plurality ofmicrofluidic channels 31 may be arranged in parallel. In otherembodiments, one or more microfluidic channels 31 may cross pathswithout being fluidly interconnected.

The microfluidic channels 31 may be formed of various materialsincluding polymers, such as silicon, glass or polymers, e.g.,polydimethylsiloxane (PDMS). In some embodiments, the microfluidicchannels 31 may be stacked on top of each other. The positioning,orientation, and arrangement of the microfluidic channels 31 may varyand thus should not be construed as limited by the exemplary figures ordescription herein.

As shown in FIG. 8, another exemplary embodiment of the fluid receivingdevice 30 may comprise one or more microwell arrays 32. Each microwellarray 32 will generally comprise an array of microwells into which thebiological fluid 16 may be transferred for scanning by the scanner 70using the various techniques discussed herein. The shape, size, anddensity of the individual wells of the microwell array 32 used herewithmay vary in different embodiments.

The number of microwell arrays 32 utilized for scanning each biologicalfluid 16, or for scanning multiple biological fluids 16, may vary indifferent embodiments. In some embodiments, a single microwell array 32may be utilized such as shown in FIG. 8. In other embodiments, multiplemicrowell arrays 32 may be utilized, such as for scanning multiplebiological fluids 16 simultaneously or in-turn.

By way of example, multiple microwell arrays 32 may be arranged inparallel so as to allow for multiple biological fluids 16, either fromthe same or different biological fluid sources 17, to be simultaneouslyscanned and then routed accordingly. In this manner, multiple types ofbiological fluids 16 from the same or multiple biological fluid sources17 may be processed simultaneously. By way of example, a firstbiological fluid 16 may be processed in a first microwell array 32 and asecond biological fluid 16 may be processed in a second microwell array32. Additional microwell arrays 32 may similarly be operated in parallelto suit any number of biological fluids 16. Some embodiments couldcombine the use of microwell arrays 32 and microfluidic channels 31.

As a further example, multiple microwell arrays 32 may be arranged inseries such that a single biological fluid 16, or multiple biologicalfluids 16, may be scanned multiple times in multiple stages. Forexample, a first microwell array 32 may process a biological fluid 16 afirst time, and then the biological fluid 16 may be transferred to asecond microwell array 32 in series with the first microwell array 32 soas to process the biological fluid 16 a second time. Subsequentmicrowell arrays 32 may also be added, allowing for multi-stageseparation of the biological fluid 16 as discussed herein. In someembodiments, multiple microwell arrays 32 may be arranged both inparallel and in series so as to allow simultaneous, multi-stageprocessing of multiple biological fluids 16.

It should also be appreciated that the type of microwell array 32 mayvary in different embodiments. The microwell array 32 may comprisesingle-use wells (e.g. parylene valves) or multi-use wells (e.g., piezolocks). The number of microwells included in each microwell array 32 mayvary in different embodiments to suit different applications. Further,the manner in which the biological fluid 16 is introduced into themicrowell array 32 may vary in different embodiments. By way of exampleand without limitation, the biological fluid 16 may be introduced intothe microwell array 32 utilizing pumps, valves, microfluidic devices,pipettes, or various combinations thereof.

As best shown in FIG. 36A, another exemplary embodiment of a fluidreceiving device 30 may comprise a droplet generator 34 in which cellsare encapsulated within droplets 39 such that the droplets 39 may bescanned by the scanner 70. Such an embodiment may rely on the use ofdroplets 39 to compartmentalize cells in nanoliter scale compartments.Similar systems have been previously used as reaction chambers fortranscriptomic analysis. Utilizing the systems and methods describedherein, cells may be encapsulated into droplets 39. Those droplets 39may then be scanned by the scanner 70 to differentiate between healthycells (e.g., normal blood cells) or unhealthy cells (e.g., cancercells). The generated droplets 39 may comprise various shapes and sizes.The generated droplets 39 may be spherical or non-spherical (e.g.,oblong).

In an embodiment utilizing droplet sorting such as shown in FIG. 36A, adefined microfluidic channel-cross design, such as a droplet generator34, may be utilized. Using the droplet generator 34, two or moreimmiscible phase channels 35, 36 will generally meet at an angle togenerate droplets 39. A first exemplary immiscible phase may comprise adispersed phase channel 35 through which an aqueous solution containingcells 18 may be routed. A second exemplary immiscible phase channel maycomprise one or more continuous phase channels 36, through which oil maybe routed.

Continuing to reference FIG. 36A, it can be seen that the dispersedphase channel 35 includes an aqueous solution containing cells 18 fromthe biological fluid 16. The continuous phase channel 36 includes oil oroils which are immiscible with the aqueous solution. The dispersed phasechannel 35 and continuous phase channel 36 meet at a juncture 37 at anangle such that droplets are generated which compartmentalize the cellsto be scanned. In the embodiment shown in FIG. 36A, multiple continuousphase channels 36 are utilized.

As can be seen in FIG. 36A, the juncture 37 may comprise a four-wayjuncture 37 having a single dispersed phase channel 35, a pair ofcontinuous phase channels 36, and a scanning channel 38 wherein thedroplets 39 containing compartmentalized cells 18 are scanned. It shouldbe appreciated, however, that such a configuration of the juncture 37and associated channels 35, 36, 38 may vary in different embodiments.For example, in some embodiments, only a single continuous phase channel36 may be utilized.

The angle at which the dispersed phase channel 35 and continuous phasechannel(s) 36 converge at the junction 37 may vary in differentembodiments. In the exemplary embodiment shown in the figures, a pair ofcontinuous phase channels 36 meets at right angles with a singledispersed phase channel 35. However, various other angles may beutilized in different embodiments to suit different applications.Further, the angle of the scanning channel 38 with respect to thedispersed phase channel 35 and continuous phase channel(s) 36 may alsovary in different embodiments.

The flow rates of the respective phase channels 35, 36 define thethroughput of the system. The flow rate within the respective phasechannels 35, 36 may vary in different embodiments. Generally, the flowrate of the dispersed phase channel 35, generally containing cells 18from the biological fluid 16 in an aqueous media, such as an aqueoussolution, will be greater than the flow rate of the continuous phasechannel 36, generally containing an oil. However, different flow ratesmay be utilized to suit different biological fluids 16, scanners 70, orother considerations. Further, the rate of flow through the scanningchannel 38 may vary in different embodiments. In some embodiments, thescanning channel 38 may include a channel lock such as an inlet valve 33so as to pause or reduce flow rate while cells 18 are being scanned.

The size of the droplet generator 34 and the ratio of the dispersedphase channel 35 when compared with the continuous phase channel 36 willgenerally define the size of generated droplets 39 exiting the juncture37. Thus, the size of the droplet generator 34, and the ratio of thesizes of the dispersed phase channel 35 and continuous phase channel 36,may vary in different embodiments to suit different sizes of generateddroplets 39 to be scanned by the scanner 70. Accordingly, the size ofthe droplet generator 34 and ratio between the respective phase channels35, 36 should not be construed as the exemplary embodiment shown in FIG.36A.

Cells 18 of the biological fluid 16 are routed first through thedispersed phase channel 35 in an aqueous media such as an aqueoussolution. The aqueous media containing the cells 18 from the biologicalfluid 16 passes through the juncture 37, at which point oil from thecontinuous phase channel(s) 36 is introduced to encapsulate the cells 18in droplets 39, with the oil being immiscible with the aqueous solutionso as to generate the droplets 39. FIG. 36A illustrates that oil isintroduced at a right angle from two separate continuous phase channels36. It should be appreciated that oil may be introduced at various otherangles, and more or less continuous phase channels 36 may be utilizedthan are shown in the exemplary embodiment shown in the figures.

In an exemplary embodiment, individual droplets 39 exit the juncture 37through a scanning channel 38 to pass a scanner 70 such as a digitalholographic microscope. The scanner 70 is configured to scan eachdroplet 39 as it passes out of the juncture 37 and through the scanningchannel 38. The scanner 70 may be configured to scan each droplet 39within the scanning channel 38 separately in-turn, or may be configuredto scan multiple droplets 39 simultaneously. In some embodiments,multiple droplets 39 may be scanned simultaneously within the scanningchannel 38 to determine if any of the droplets 39 encapsulate cells 18that are undesirable or malignant; with the droplets 39 being sorted bydiverting the flow of the droplets 39 according to the scan results.

The results from the scanner 70 are then analyzed, such as by a controlunit 80, to determine if the encapsulated cells 18 within each droplet39 are healthy or unhealthy (e.g., malignant). If the cells 18 arehealthy, those cells 18 may be separated from other cells 18 (e.g.,malignant cells) by use of one of a pair of valves 40. In the exemplaryembodiment shown in FIG. 36A, it can be seen that a first valve 40 leadsto a return path 60 for healthy cells and a second valve 40 leads to anisolation path 50 for all other cells 18 (unidentified cells 18 ormalignant cells 18).

When a healthy cell 18 is scanned by the scanner 70, the first valve 40is opened and the second valve 40 is closed. When an undesirable ormalignant cell 18 is scanned by the scanner 70, the second valve 40 isopened and the first valve 40 is closed. The first valve 40 may lead toa return path 60 such that healthy cells 18 may be returned to thepatient 12 for therapeutic purposes, or may lead to an isolation path 50to be sequestered for diagnostic purposes. The second valve 40 may leadto an isolation path 50 to be sequestered.

It should also be appreciated that the fluid receiving device 30 maycomprise one or more inlet valves 33 such as channel locks adapted topause flow of the biological fluid 16 into of the fluid receiving device30. Such inlet valves 33 may comprise valves, locks, or other structureswhich are adapted to block flow of the biological fluid 16 at certaintimes. For example, one or more inlet valves 33 of the fluid receivingdevice 30 may be engaged to pause flow of the biological fluid 16 intothe fluid receiving device 30 while scanning is being performed by thescanner 70.

D. Scanning Techniques

As described and shown herein, a wide range of scanning techniques maybe utilized to scan a biological fluid 16 and identify any constituentbiological elements thereof, such as but not limited to white bloodcells, red blood cells, CTCs, platelets, host cells, cell-free plasma,pathogens, and the like. The types of biological fluids 16 that can bescanned include a wide range of biological fluids 16, such as but notlimited to blood, lymphatic fluid, cerebrospinal fluid, sweat, urine,pericardial fluid, stools, saliva, and the like. In some embodiments,multiple types of biological fluids 16 may be scanned either together orin turn. For example, both blood and cerebrospinal fluids may besimultaneously or sequentially scanned by the same fluid receivingdevice 30.

Exemplary techniques for scanning may include Phase contrast microscopy(PCM), DIC Microscopy, Hoffman modulation, polarized light microscopy,digital holographic microscopy, confocal scanning optic microscopy(CSOM), or laser scanning optic microscopy (SOM) to measure voxelfluorescence, bright-field microscopy, dark-field illumination, Ramanspectrometry to measure Raman Scattering, Optical interferometry tomeasure optical interference, total internal reflection fluorescencemicroscopy to measure evanescent effect, planar waveguides forrefractive index detection, photonic crystal biosensors for measure ofbiomolecules on cell surfaces, and light property modulation detectionssuch as surface plasmon resonance (SPR) detection.

With respect to digital holographic microscopy, digital holography isused to record a wave front diffracted from an object by a light source72. Utilizing the interference of light from the light source 72, bothamplitude and phase information of an object wave 77 may be recorded toproduce a hologram containing the information of the object wave 77. Athree-dimensional image may then be reconstructed from the hologram bythe control unit 80.

In an embodiment utilizing digital holographic microscopy, the scanner70 may include a light source 72 as previously discussed. The lightsource 72 may comprise various types of illuminating devices, such asbut not limited to a laser such as a monochromatic laser. A pair oflaser light waves is generated from the light source 72 by dividing thelaser beam with a beam splitter 74 such that one of the split lightwaves illuminates the biological fluid 16. The light diffracted from thebiological fluid 16 forms an object wave 77, which illuminates thescanner 70 and is collected by the microscope objective 73. Theremaining laser light wave is directly detected by the microscopeobjective 73 of the scanner 70 to serve as a reference wave 78. Theobject and reference waves 77, 78 interfere with each other at thescanner 70 to form an interference fringe image which is scanned by animage sensor 76.

Continuing to reference digital holographic microscopy, the object andreference wave 77, 78 fronts may be joined by the beam splitter 74 suchthat the object and reference wave 77, 78 fronts interfere and create ahologram which is detected by an image sensor 76. The control unit 80may then process the digital hologram, with the control unit 80functioning as a digital lens to calculate a viewable image of theobject wave 77 front utilizing a numerical reconstruction algorithm.

While a microscope objective 73 may be used to collect the object wave77 front, it should be appreciated that the microscope objective 73 isonly used to collect light waves and not to form an image. Thus, themicroscope objective 73 may comprise a simple lens, or may be omittedentirely in some embodiments. The interference pattern (hologram) maythus be recorded in such embodiments by a digital image sensor 76.

Digital holographic microscopy may be utilized to observe living cellswithin the biological fluid 16. From the recorded interference patternof such living cells, the intensity and phase shift across variouspoints of the cells may be numerically computed by the control unit 80.The control unit 80 may thus measure the phase delay images ofbiological cells within the biological fluid 16 to provide quantitativeinformation about the morphological properties (e.g., cellular dry mass,surface texture, shape, etc.) of individual cells within the biologicalfluid 16.

In an exemplary embodiment, the systems and methods described herein mayutilize these quantitative indicators of morphological properties in analgorithm to distinguish between cell types within the biological fluid16. By way of example and without limitation, the control unit 80 may beadapted to extract parameters such as cell thickness, cell area, cellvolume, cell dry mass, the phase shift across the cell, surfaceroughness and texture, cell shape, elongation, convexity, luminance,circularity, solidity, and the like.

Various types of digital holography may be utilized with the systems andmethods described herein, including but not limited to off-axis Fresnel,Fourier, image plane, in-line, Gabor, and phase-shifting digitalholography. FIG. 31 illustrates an off-axis embodiment. By utilizingdigital holographic microscopy, the control unit 80 may differentiatebetween the various constituents 13 within a biological fluid 16 samplefor further processing utilizing the systems and methods describedherein.

It should be appreciated that multiple laser wavelengths may be utilizedwhen scanning the biological fluid 16 with digital holographicmicroscopy. It has been shown that the refraction amount increases asthe wavelength of light decreases. Thus, shorter wavelengths of light(e.g., violet and blue) are more slowed and consequently experience morebending than longer wavelengths of light (e.g., orange and red).

Since the morphological parameters in digital holographic microscopy aredependent upon the wavelength of the laser used, an exemplary embodimentof a scanning technique relying upon digital holographic microscopy mayutilize multiple lasers each having different wavelengths. By way ofexample, the analysis may be initially conducted using a light source ata first wavelength. If the sample of cells requires additionalconfirmation, the light source may be switched to a differentwavelength.

E. Ex Vivo Testing

The systems and methods described herein may be utilized for ex vivotesting of various drugs and treatments on a patient 12 specific basis.While established tissue culture cell lines are often used for in vitrodrug sensitivity assays, such cell lines are not truly representative ofthe cellular heterogeneity evidenced during metastasis and recurrence inspecific patients 12. Thus, it is would be far more desirable to testsuch drugs and treatments on patient-derived CTCs and CTC-clusters.

Patient-derived CTCs and CTC-clusters offer greater precision inpredicting outcomes for a particular patient 12, as they represent theheterogeneity profile for that particular patient 12 at that particulartime. Critically, patient-derived CTCs and CTC-clusters may exhibitenhanced resistance to chemotherapy during stages of relapse.

Using the systems and methods described herein, sufficient CTCs andCTC-clusters may be extracted to permit both genomic and transcriptomicprofiling. This allows for “direct-to-drug” ex vivo testing oftreatments and drug agents to identify an optimal course of therapy fora particular patient 12 at any particular time during the progress oftreatment.

FIG. 34 illustrates an exemplary embodiment in which a portion of CTCswhich have been identified and separated by the systems and methodsdescribed herein may be profiled for heterogeneity. As can be seen, CTCsand CTC-clusters may undergo genomic and transcriptomic profiling todetermine potentially relevant drugs or treatments. After identifyingpotential drugs or treatments based upon the isolated CTCs andCTC-clusters from a particular patient 12, relevant drug targets may beidentified. The drugs or treatments may then be tested on thatparticular patient's cancer cells ex vivo to identify optimal pathways.

In this manner, drugs and treatments may be optimized for each patient12 based upon the cancer cells identified and separated by the methodsand systems described herein to allow for precision drug selection thatis unique to each patient 12 in consideration of the cancer cells uniqueto that patient 12 and in consideration of the course of treatment tothat point for that patient 12.

F. Multi-Stage Separation

As previously discussed, the systems and methods described herein may beutilized for multi-stage separation of non-healthy or malignant cellsfrom healthy cells. FIG. 35 illustrates an exemplary embodiment of sucha multi-stage separation system in which multiple stages are utilized toseparate CTCs and CTC-clusters from a biological fluid 16. FIG. 36Billustrates another exemplary embodiment in which an isolation path 50is split into additional channels for repeated, additional enrichmentthrough scanning.

As shown in FIG. 35, the first stage of the multi-stage separationmethodology involves receiving a biological fluid 16 from the patient12. The biological fluid 16 may enter the multi-stage separationdirectly from the patient 12. At the first stage, passive inertialsorting, with or without the use of buffer fluids, may be utilized toseparate red blood cells and CTC-clusters from the biological fluid 16.Red blood cells separated at this first stage may be returned to thebiological fluid source 17 and CTC-clusters may be isolated for furtherprocessing.

Continuing to reference FIG. 35, CTC-clusters and red blood cells areseparated from the biological fluid 16 in the first stage by inertialsorting. The remaining cells, which will generally include additionalred blood cells, white blood cells, and CTCs, will then be passed ontothe second stage. Such remaining cells may be passed onto a fluidreceiving device 30 for a high-throughput optical sort utilizing neuralnetwork algorithms. The second stage will thus be utilized to furtherseparate the cells of the biological fluid 16. Any separated red andwhite blood cells may then be returned to the patient 12, withidentified CTCs and any remaining white blood cells being passed on to athird stage.

As shown in FIG. 35, the third stage may comprise precision opticalsorting through use of an additional fluid receiving device 30 that isin series with the fluid receiving device 30 of the second stage.Interpretable machine learning classifiers may be utilized to separatethe remaining white blood cells, thus leaving only CTCs which areseparated at the third stage. In this manner, CTC-clusters and CTCs maybe individually separated from a biological fluid 16, with white and redblood cells from the biological fluid 16 being returned to the patient12. The individually separated CTC-clusters and CTCs may then beretained for further processing, such as ex vivo testing, therapeutics,or diagnostics. Additional stages may also be added as-needed forfurther enrichment.

G. Optical Filtration of Subsets of Healthy Cells

The methods and systems described herein may be utilized to filterspecific subsets of healthy cells (e.g., T-cells) from other types ofcells. In such an embodiment, the library of desirable constituents 15(e.g., healthy cells) may be configured to exclude specific subsets ofhealthy cells (e.g., T-cells) so as to allow the biological fluidfiltration system 10 to filter such subtypes of healthy cells. Aspreviously indicated, such methods may be applied for human orveterinary uses. The biological fluid 16 may be scanned for subsets ofhealthy cells utilizing fluid receiving devices 30 such as but notlimited to microfluidic channels 31, microwell arrays 32, and dropletgenerators 34.

In the course of some therapies and treatments, such as Chimeric antigenreceptor (CAR) T-cell therapy, T-cells engineered with chimeric antigenreceptors may be utilized for cancer therapy. As is known in the art,T-cells are a type of white blood cell which develops in the thymusgland and play a central role in the body's immune response. CAR-Timmunotherapy is utilized to modify T-cells to recognize cancer cells inorder to more effectively target and destroy them.

T-cells are harvested, genetically altered, and then the resulting CAR-Tcells are infused into the patient 12 to attack cancer cells. Such CAR-Tcells may be derived from T-cells in the patient's 12 own blood(autologous) or derived from the T-cells of another healthy donor(allogenic). Once isolated, these T-cells are genetically engineered toexpress a specific CAR, which programs the T-cells to target an antigenthat is present on the surface of tumors. In other therapies, otherforms of white blood cells (such as natural killer or “NK” cells) aresimilarly engineered to fight cancers.

In an exemplary embodiment of a biological fluid filtration system 10for filtration of subsets of healthy cells, the biological fluid source17 may be an aphaeretic extract from a therapeutic apheresis orleukapheresis machine containing white blood cells including T-cells.The systems and methods described herein may be utilized to extract justthe T-cells by recognizing all healthy cells except for the T-cells, andthen filtering out the undesirable cells (the T-cells). As a furtherexample, other types of healthy cells may be omitted from recognition,such as NK cells, so that those healthy cells may be filtered out in asimilar manner.

FIG. 37 illustrates an exemplary method of filtering out T-cells from abiological fluid 16 containing other types of healthy cells. It shouldbe appreciated that the biological fluid source 17 may be human orveterinary and may include such biological fluids 16 as blood, lymphaticfluid, cerebrospinal fluid, sweat, urine, pericardial fluid, stools, andsaliva.

As shown in FIG. 37, an optional pre-filtration session may be conductedto create a reference data 91 in which a sample of biological fluid 16,from the patient 12 or others, is pre-processed by optically scanningthe sample with a scanner 70 and recognizing certain biological fluidconstituents 13 such as certain types of cells within the biologicalfluid 16 using an algorithm run by a control unit 80. The optical scanof the subsample allows for a software algorithm run by the control unit80 to analyze the scanned data from the subsample and recognize anysubsets of healthy blood cells (e.g., red blood cells, white bloodcells) while not recognizing certain subsets of healthy blood cells(e.g. T-cells). Such pre-processing may be utilized to create a controlgroup for recognizing different types of cells during the processingphase.

Continuing to reference FIG. 37, it can be seen that in an exemplaryembodiment a patient 12 may undergo leukapheresis. The leukapheresisextract from the patient containing healthy cells is channeled through afluid receiving device 30 such as microfluidic channels, microwellarrays, or droplet generators and the extract is scanned by the scanner70. The control unit 80 then performs image processing and detection ofthe contents within the fluid receiving device 30, including use of anypre-processing findings if pre-processing had previously occurred.

The control unit 80 will analyze the results of the scan from thescanner 70 to determine where to route the biological fluid 16. If cellsother than recognized healthy cells are present, the biological fluid 16may be routed along a reprocessing path 62 for the contents to beprocessed for extraction of any undesirable cells (e.g., T-cells). Ifonly recognized healthy cells are present, the contents may be routedalong an isolation path 50 to be discarded or retained for further usewithout cell extraction.

H. Presorting of Biological Fluids

FIG. 38 illustrates an exemplary method of presorting and then opticallyfiltering a portion of a biological fluid 16 of biological fluidconstituents 13 such as pathogens or CTCs. As shown in FIG. 38, anoptional pre-filtration session may be conducted to create a referencedata 91 in which a sample of biological fluid 16, from the patient 12 orothers, is pre-processed by optically scanning the sample with a scanner70 and recognizing the biological fluid constituents 13 such as cellswithin the biological fluid 16 using an algorithm run by a control unit80.

Continuing to reference FIG. 38, it can be seen that biological fluid 16is first drawn from the biological fluid source 17 during a filtrationsession. The biological fluid 16 may comprise various fluids asdiscussed herein, such as but not limited to blood, lymphatic fluid,CSF, sweat, urine, pericardial fluid, stools, and saliva. The biologicalfluid source 17 may comprise a human or an animal. After the biologicalfluid 16 is drawn from the biological fluid source 17, the biologicalfluid 16 may be presorted, such as by using a microfluidic separationmodule 100.

The presorting of the biological fluid 16 may be utilized to separatedesirable constituents 15 from the biological fluid 16 prior to furtherprocessing. The manner of presorting utilized by the microfluidicseparate module 100 may vary in different embodiments, and may includewithout limitation the use of inertial sorting, centrifugal sorting,microfluidic sorting, and the like. Characteristics utilized duringpresorting may include size, density, inertial hydrodynamic, antigenbinding affinity, motility, centrifugation, electrical charge, electricdipole moment, or magnetism.

The presorting of the biological fluid 16 by the microfluidic sortingmodule 100 allows for the initial separation of biological fluidconstituents 13 without optical scanning. Any such biological fluidconstituents 13, such as CTC, may be immediately routed along a returnpath 60 back to the biological fluid source 17, or may be sequesteredalong an isolation path 50 for further processing. After the presortingstep is completed, a mixture of undesirable constituents 14 anddesirable constituents 15 may then be transferred to a fluid receivingdevice 30 for further processing.

Continuing to reference FIG. 38, it can be seen the mixture ofundesirable constituents 14 and desirable constituents 15 is opticallyscanned on the fluid receiving device 30 by a scanner 70. The controlunit 80 then performs image processing and detection to identify thecontents and differentiate between undesirable constituents 14 anddesirable constituents 15. If undesirable constituents 14 are detectedin the sample, the sample is not returned to the biological fluid source17, but may instead be sent along an isolation path 50 for sequestrationand optional diagnostics of the contents to identify pathogens, CTC,CTC-clusters, host cells, cell free plasma, and the like.

Any samples comprising only undesirable constituents 14 after thepresorting stage may be immediately directed along an isolation path 50and not returned to the biological fluid source 17. Such samples may besequestered along the isolation path 50 for optional diagnostics and/ortherapeutics as discussed herein.

FIG. 39 illustrates another exemplary method of presorting and thenoptically filtering a portion of a biological fluid 16 which, byutilizing multiple outlet ports, may be utilized to sequester andseparately hold isolated biological fluid constituents 13 such as CTCs,CTC-clusters, white blood cells, cell-free plasma, and the like. Thesequestered outputs may then be reprocessed single or multiple timesusing the systems and methods described herein, without the aphaereticcomponents, to further enrich the isolation of such biological fluidconstituents 13.

As shown in FIG. 39, an optional pre-filtration session may be conductedto create a reference data 91 in which a sample of biological fluid 16,from the patient 12 or others, is pre-processed by optically scanningthe sample with a scanner 70 and recognizing the biological fluidconstituents 13 such as cells within the biological fluid 16 using analgorithm run by a control unit 80. Biological fluid 16 is first drawnfrom the patient 12. If necessary, an anti-coagulant may be applied tothe biological fluid. A fluid pump may draw the biological fluid 16through a fluid pressure sensor and pre-filter pressure sensor prior toentering a pre-sorting module. The pre-sorting module utilizes thetechniques and/or characteristics described herein, such as but notlimited to inertial, size-based, centrifugal, dielectric, and/oracoustic to separate biological fluid constituents 13 from thebiological fluid 16. In some embodiments, the use of additional buffersor dilution liquids or reagents may be employed in the pre-sortingmodule. In such examples, the embodiment might include ports andreservoirs to insert, collect, and/or replenish such fluids.

A first set of biological fluid constituents 13, such as red bloodcells, plasma, and small cells, may be transferred to a fluid chamber totemporarily hold filtered plasma and healthy cells. Separated largeCTC-clusters may be retained and sequestered for further processing suchas diagnostics. Separated white blood cells and small CTCs may betransferred into one or more fluid receiving devices 30 such asmicrofluidic channels 31 arranged in parallel. Each fluid receivingdevice 30 is independently scanned, and its contents then undergo imageprocessing and content detection by the control unit 80, which mayutilize findings from any optional pre-filtration session.

If undesirable constituents 14 such as cells other than healthy cellsare detected, the contents may be re-routed back to the fluid receivingdevices 30 by a reprocessing path 62 for further enrichment of CTCs. Ifno cells other than healthy cells are detected, the contents may becombined with the filtered plasma and healthy cells, pass through afluid pump and pressure sensor, undergo air bubble removal, and returnedto the biological fluid source 17 by the return path 60.

FIG. 40 illustrates yet another method of presorting and then opticallyfiltering a portion of a biological fluid 16. Such embodiments mayinclude the use of diagnostic systems to separate pathogens or CTCs frombiological fluid 16 samples for downstream diagnostic (e.g., genomic,transcriptomic, metabolomics, drug sensitive, drug resistance, etc.)characterization. FIG. 40 illustrates such a diagnostic embodimentwithout a return path 60 back to the biological fluid source 17.

As shown in FIG. 40, an optional pre-filtration session may be conductedto create a reference data 91 in which a sample of biological fluid 16,from the patient 12 or others, is pre-processed by optically scanningthe sample with a scanner 70 and recognizing the biological fluidconstituents 13 such as cells within the biological fluid 16 using analgorithm run by a control unit 80. At the filtration session,biological fluid 16 is drawn from the source and entered into apre-sorting module to separate biological fluid constituents 13 (e.g.,inertial, size-based, centrifuge, dielectric, acoustic, etc.). Anyseparated red blood cells, plasma, and small cells are transferred to afluid chamber to hold plasma and healthy cells. Any separated largeCTC-clusters may be sequestered for further processing.

Samples including separated white blood cells and small CTCs may betransferred by inertial focusing into one or more parallel fluidreceiving devices 30. Fluid flow is temporarily halted, and each fluidreceiving device 30 is scanned by a scanner 70. The control unit 80conducts image processing and content detection of each such samplein-turn.

If cells other than healthy cells are present, such as extracted CTCsalong with potentially some healthy cells, the sample may be reroutedback to the fluid receiving devices 30 by a reprocessing path 62 forfurther enrichment of CTCs. Enriched contents may be routed along anisolation path 50 for diagnostic processing (e.g., genomic,transcriptomic, metabolomics, drug sensitivity, drug resistance, etc.)along with any previously-separated red blood cells, plasma, smallcells, and separated large CTC-clusters.

FIG. 41 illustrates another embodiment of a method of presorting andthen optically filtering a portion of a biological fluid 16, withhealthy cells being returned back to the biological fluid source 17. Anoptional pre-filtration session may be conducted to create a referencedata 91 in which a sample of biological fluid 16, from the patient 12 orothers, is pre-processed by optically scanning the sample with a scanner70 and recognizing the biological fluid constituents 13 such as cellswithin the biological fluid 16 using an algorithm run by a control unit80.

During the filtration session, biological fluid 16 is drawn from thebiological fluid source 17. Anti-coagulants may be applied if necessaryto prevent clotting within the system. The biological fluid 16 is thenpumped by a fluid pump past a fluid pressure sensor and a pre-filterpressure sensor. The biological fluid 16 then enters the pre-sortingmodule to separate biological fluid constituents 13 in any of themanners previously discussed. As with previous embodiments, healthycells such as separated red blood cells, plasma, and small cells may betransferred to a fluid chamber to be temporarily held. Malignant cellssuch as separated large CTC-clusters may be transferred along anisolation path 50 to be sequestered for diagnostic processing.

Samples with both healthy and malignant cells, such as separated whiteblood cells and small CTCs, may be inertial focused into one or moreparallel fluid receiving devices 30 such as microfluidic channels 31.Once within the one or more fluid receiving devices 30, such sampleswill be held in the fluid receiving device 30 as optical scans areperformed by one or more scanners 70. In an embodiment in which multiplescanners 70 are utilized, each of the samples may be scannedsimultaneously. In an embodiment in which a single scanner 70 isutilized, each of the samples may be held in place while each sample issequentially scanned.

It should also be appreciated, with this embodiment and with the othersdescribed herein, that a single fluid receiving device 30 may beutilized to scan multiple distinct samples at different times. In suchembodiments, a first sample will be transferred to the fluid receivingdevice 30, scanned, and the results will be transferred to the controlunit 80. Upon completion of analysis of that sample, fluid flow may beresumed to transfer another sample onto the fluid receiving device 30for scanning. These steps may be repeated until all of the samples havebeen scanned.

The results of each optical scan (e.g., images or other cellcharacteristics) are transferred to the control unit 80 for imageprocessing and content detection. This step may be repeated until all ofthe samples with both healthy and malignant cells have been scanned andprocessed as described above. If healthy cells are detected amongunhealthy cells (e.g., CTCs), the sample may be returned to the fluidreceiving device 30 along a reprocessing path 62 for further scanningand processing. Alternatively or after multiple such scans andenrichment, the sample may be sequestered for diagnostic processingusing any of the methods previously discussed herein.

If only healthy cells are detected in a sample (e.g., only white bloodcells and no CTCs), the sample may be transferred via a fluid pump andpressure sensor back to the patient 12 or biological fluid source 17along a return path 60. In some embodiments, any filtered plasma andhealthy cells in the fluid chamber may similarly be returned to thebiological fluid source 17, either with the white blood cells orseparately. Air bubbles will also be removed prior to return to thebiological fluid source 17. In this manner, the embodiment shown in FIG.41 allows for only confirmed healthy cells to be returned to the patient12, with any other cells (e.g., malignant cells or undesirable cells)being sequestered for disposal or for diagnostic processing.

In another exemplary embodiment such as shown in FIG. 42, non-aphaereticmethods may be utilized for the removal of tumor cell contaminants fromautologous stem-cell transplant products. Autologous stem celltransplants are generally used in patients 12 who need to undergo highdoses of chemotherapy and radiation to cure their disease. Thesetreatments can be toxic and thus damage the bone marrow. An autologousstem cell transplant helps to replace the damaged bone marrow, but it isoften reported that the process to collect stem cells from the patientmay lead to contamination of such products with tumor cells. Using themethod shown in FIG. 42, the biological fluid filtration system 10 maybe used to prevent or remove such contamination of stem cells.

As shown in FIG. 42, a pre-filtration session may be conducted torecognize healthy cells. During the filtration session, biological fluid16 such as leukapheresis product from the biological fluid source 17 istransferred to a presorting module to separate fluid constituents by anyof the methods described herein. As with the other embodiments, thepre-sorting may separate healthy cells such as red blood cells, plasma,and small cells which are transferred to a fluid chamber fortransplantation. Malignant cells such as large CTC-clusters may besequestered for diagnostic processing using any of the methodspreviously described.

With respect to the remaining samples containing both healthy andmalignant or undesirable cells, inertial focusing may be utilized totransfer such samples onto one or more fluid receiving devices 30. Eachsample is then scanned (either simultaneously using multiple fluidreceiving devices 30 in parallel or sequentially using a single fluidreceiving device 30 or multiple fluid receiving devices 30 in series)and the results transferred to the control unit 80 for image processingand content detection.

If cells other than healthy cells are detected, such samples may bererouted back to the fluid receiving device 30 along a reprocessing path62 for further processing or may be sequestered for diagnosticprocessing along an isolation path 50 using any of the methods describedherein. If only healthy cells are detected, such samples may betransferred to the fluid chamber with the healthy cells from thepre-sorting to be transplanted back to the patient 12.

FIG. 43 illustrates an embodiment in which multiple pre-sortingtechniques are utilized in conjunction with optical filtration in abiological fluid filtration system 10. Biological fluids 16 are drawnfrom a biological fluid source 17 such as an animal or human. A passiveinertial sort may be utilized to separate any CTC-clusters which may besequestered for disposal or diagnostics. After the initial inertialsort, centrifugation may be utilized to remove white blood cells andCTCs (e.g., leukapheresis).

During leukapheresis, some red blood cells and plasma may be returned tothe patient 12. The leukapheresis product containing white blood cellsand CTCs may be transferred to a third stage for optical filtrationusing the methods described herein to separate the CTCs for diagnosticprocessing. Thus, the biological fluid 16 is pre-filtered acrossmultiple stages prior to optical filtration. As a further example, bloodcould first be filtered using microfluidic inertial mechanisms to sortout large CTC-clusters. The remainder undergoes therapeutic apheresis toseparate white blood cells and CTCs. That mix of cells is then filteredusing the optical filtration techniques discussed herein.

I. Body-Worn Aphaeretic Optical Filtration Device

While the various embodiments previously discussed are typicallyconducted in clinical or lab settings, certain embodiments may allow forthe systems and methods described herein to be utilized in a body-wornconfiguration outside of a clinical or lab setting.

FIGS. 44A, 44B, and 45 illustrate an exemplary embodiment of such abody-worn device 120. The body-worn device 120 may utilize opticalcomponents, including one or more fluid receiving devices 30 and one ormore scanners 70, to filter undesirable constituents 14 from variousbiological fluids 16, such as blood, lymphatic fluid, cerebrospinalfluid, sweat, urine, pericardial fluid, stools, and saliva. Thebiological fluid source 17 may be veterinary or human.

The body-worn device 120 will generally comprise a housing 121 whichhouses all of the power, optical, microfluidic, computational, andcommunication components necessary for the patient 12 utilizing thebody-worn device 120 to freely move about. The body-worn device 120 willgenerally include a power source 122 for powering the various componentsof the body-worn device 120. The power source 122 may comprise varioustypes of batteries, including disposable and rechargeable batteries. Insome embodiments, the power source 122 may comprise solar cells.

The body-worn device 120 may also include various indicators 132 adaptedto convey various information to the user of the body-worn device 120.For example, indicators 132 may be utilized to indicate the power status(on or off) of the body-worn device 120, the charge remaining in thepower source 122, the remaining volume available in the cartridge 126(e.g., whether the cartridge is full or nearing full), and the like. Theindicators 132 may be visual (e.g., lights) and/or audible (e.g.,alarms). For example, an audible and/or visual alarm may be activatedwhen the cartridge 126 is nearing full, or if the power source 122 isrunning out of charge.

FIG. 44B illustrates an exemplary embodiment of a body-worn device 120which includes an anti-coagulant insert 134 which includesanti-coagulant which is applied to the biological fluid 16 within thebody-worn device 120 to prevent clotting or coagulation of thebiological fluid 16 within the body-worn device 120 as discussed herein.A removable anti-coagulant insert 134 of such anti-coagulant may beremovably connected to the body-worn device 120 such that theanti-coagulant within the anti-coagulant insert 134 may be easilyreplenished as-needed. Also shown in FIG. 44B is a buffer fluid insert136 which may be utilized to collect or replenish optional bufferfluids. Such buffer fluids such as dilution fluids may be utilized forpre-sorting within the body-worn device 120 as discussed herein.

As shown in FIG. 45, the body-worn device 120 will generally comprise aninlet 123 and an outlet 124. The inlet 123 and outlet 124 of thebody-worn device 120 are generally adapted to be connected tointravenous (IV) or catheter ports 129 a, 129 b of a catheter 128 whichis inserted into the body of the patient 12 to receive a biologicalfluid 16 from a biological fluid source 17. It should be appreciatedthat the catheter 128 may be installed at various locations on apatient's 12 body, and thus FIGS. 44A and 44B should not be construed aslimiting in that regard. The body-worn device 120 may be worn adjacentto the catheter 128 entering the body, or more distant from thatposition. Thus, the length of the catheter 128 may vary widely indifferent embodiments.

In some embodiments, the catheter 128 may be fluidly connected to thevascular system of a patient 12 such that the vascular system of thepatient 12 acts as a biological fluid source 17 for a biological fluid16 comprised of blood. In other embodiments, the catheter 128 may befluidly connected to the nervous system of a patient 12 such that thenervous system of the patient 12 acts as a biological fluid source 17for a biological fluid 16 comprised of cerebrospinal fluid. In otherembodiments, the catheter 128 may be fluidly connected to the lymphaticsystem of a patient 12 such that the lymphatic system of the patient 12acts as a biological fluid source 17 for a biological fluid 16 comprisedof lymphatic fluid.

By way of example, biological fluid 16 comprised of blood may beaccessed through arteriovenous grafts, fistulas, or catheters 128commonly used in aphaeretic treatments such as dialysis. Access to othertypes of biological fluid 16 such as cerebrospinal fluid could bethrough the lumbar, peritoneum, or the ventricles in the skull. Thus, itshould be appreciated that the catheter 128 may be positioned at variouslocations on the body of the patient 12, such as but not limited to thehead, arms, chest, legs, hands, feet, and the like.

Continuing to reference FIG. 45, it can be seen that the body-worndevice 120 may comprise one or more pumps 125 for controlling flow ofthe biological fluid 16 entering and exiting the body-worn device 120.The pump 125 may include a pressure sensor for monitoring the pressureof the biological fluid 16. The pump 125 may be configured to runcontinuously or only at certain times or depending upon certainconditions. The pump 125 may be operated by the control unit 80.

As an example, the pump 125 may activate to draw a sample comprising aset volume of biological fluid 16 onto the fluid receiving device 30.Upon the set volume of biological fluid 16 being transferred to thefluid receiving device 30, the pump 125 may deactivate during thescanning process. After scanning the sample of the biological fluid 16,the pump 125 may activate again so as to direct the scanned sample ofthe biological fluid 16 either to a removable cartridge 126 or back tothe biological fluid source 17 via the outlet 124. In some embodiments,the pump 125 may be adapted to automatically deactivate under certainconditions, such as upon pressure being detected as being above or belowcertain thresholds.

The biological fluid 16 will generally be routed from the inlet 123 intoa microfluidic fluid receiving device 30 such as is described herein. Itshould be appreciated that multiple fluid receiving devices 30 may beutilized in the body-worn device 120 in certain embodiments, either inparallel or in series. When the biological fluid 16 is in the fluidreceiving device 30, the scanner 70 will scan the contents of the fluidreceiving device 30. An anti-coagulant may be applied to the biologicalfluid 16 prior to entering the fluid receiving device 30 so as toprevent the biological fluid 16 from coagulating while in the body-worndevice 120.

One or more scanners 70 are directed towards the fluid receiving device30 to scan the biological fluid 16 within the fluid receiving device 30.The scanner 70 will generally be communicatively connected to a controlunit 80. The control unit 80 may be located within the housing 121 ofthe body-worn device 120, such as by use of a microprocessor,microcontroller, system-on-a-chip, or the like. In some embodiments, thescanner 70 may be communicatively connected to a remote control unit 80,such as through use of a communications network. By way of example, thescanner 70 may be communicatively connected to a control unit 80 byBluetooth, Wi-Fi, radio waves, or various other communications methodsknown in the art.

The results of the optical scan of the biological fluid 16 within thebody-worn device 120 by the scanner 70 are generally transferred to thecontrol unit 80 for processing and detection of the biological fluidconstituents 13. The data from the scanner 70 is compared to thereference data 91 and analyzed by the control unit 80 to classify thebiological fluid constituents 13 within the biological fluid 16. One ormore valves 127 may be utilized to direct the biological fluid 16 alongat least two different paths depending upon the analysis of thebiological fluid 16 by the control unit 80.

If only healthy, desirable constituents 15 are detected, the one or morevalves 127 may direct the biological fluid 16 along a return path 60 tobe returned to the biological fluid source 17. More specifically, thebiological fluid 16 may exit the body-worn device 120 through its outlet124 and returned to the biological fluid source 17 by the catheter 128.

If there are undesired constituents 14 or undesired constituents such asmalignant cells, the one or more valves 127 may direct the biologicalfluid 16 along an isolation path 50 to a cartridge 126. The cartridge126 may be removable and, in some embodiments, may be disposable. Thecartridge 126 generally includes a cavity within which the biologicalfluid 16 may be stored and sequestered.

The cartridge 126 is generally removably connected to the housing 121 ofthe body-worn device 120. However, in some embodiments, the cartridge126 may instead be fixedly connected to the housing 121 and insteadinclude an access port through which the biological fluid 16 may bedrained from the cartridge 126 as-needed. Thus, the valve(s) 127 of thebody-worn device 120 will generally have one inlet and a pair ofoutlets. The inlet of the valve(s) 127 is fluidly connected to the fluidreceiving device 30. A first outlet of the valve(s) 127 is fluidlyconnected to the cartridge 126 and a second outlet of the valve(s) 127is fluidly connected to the outlet 124 of the body-worn device 120.

The cartridge 126, which may be replaceable and/or disposable, mayinclude built-in sensors to identify the protein expression of thecells. Data from such built-in sensors may be transferred to the controlunit 80 for further processing. Thus, the filtered contents of thecartridge may be diagnostically profiled by the control unit 80.

FIG. 46 illustrates an exemplary method of filtering a biological fluid16 utilizing a body-worn device 120. Prior to filtration, the catheter128 will generally be inserted within the patient 12 and fluidlyconnected to the biological fluid source 17. The catheter 128 will befluidly connected to the inlet 123 of the body-worn device 120 and, insome embodiments, to the outlet 124 of the body-worn device 120. Thus,the catheter 128 may include a pair of intravenous (IV) or catheterports 129 a, 129 b, such as an outlet catheter port 129 a and an inletcatheter port 129 b. In such embodiments, the outlet catheter port 129 ais fluidly connected to the outlet 124 of the catheter 128 and the inletcatheter port 129 b is fluidly connected to the inlet 123 of thecatheter 128 such as shown in FIGS. 44A and 44B.

Biological fluid 16 is drawn from the biological fluid source 17 throughthe catheter 128, entering the body-worn device 120 through its inlet123. Anti-coagulant may be applied to the biological fluid 16 as-neededfrom the anti-coagulant insert 134. The biological fluid 16 is drawninto the fluid receiving device 30 by the pump 125, which may becontrolled by the control unit 80. The scanner 70 then scans thebiological fluid 16 within the fluid receiving device 30, and theresulting data is transferred to the control unit 80 for analysis anddetection of the biological fluid constituents 13 within the biologicalfluid 16.

The manner by which the biological fluid 16 is scanned by the scanner 70and analyzed by the control unit 80 may vary as described herein. If apre-filtration session was performed, the data collected therefrom maybe utilized by the control unit 80 in detecting and identifying thebiological fluid constituents 13 of the biological fluid 16.

In some embodiments, the control unit 80 may be configured to maintain acount of detected biological fluid constituents 13, including healthycells, malignant cells, and unidentified cells. The control unit 80 mayalso be configured to keep track of the total count of cells analyzed,the total volume of biological fluid 16 analyzed, cells detected pervolume of biological fluid 16, and other data.

Any biological fluid 16 samples scanned by the scanner 70 and determinedby the control unit 80 to include undesirable constituents 14 aredirected by the valve(s) 127 along an isolation path 50 into thecartridge 126. Any biological fluid 16 samples scanned by the scanner 70and determined by the control unit 80 to include only healthy, desirableconstituents 15 is instead routed by the valve(s) 127 along a returnpath 60 out of the body-worn device 120 via its outlet 124 to bereturned to the biological fluid source 17 by the catheter 128.

FIG. 47 illustrates a method of filtering a biological fluid 16 with abody-worn device 120 including a presorting stage. As shown in FIG. 47,the biological fluid 16 is drawn from the biological fluid source 17into the body-worn device 120 via its inlet 123 to enter a pre-sortingstage. The presorting stage sorts the biological fluid 16 by variousmethods described herein (e.g., inertial, size, density, immunoaffinity,magnetic, dielectric). In some embodiments, the presorting stage mayinclude a miniature centrifuge within the body-worn device 120. In someembodiments, the presorting stage may utilize a buffer fluid insert 136from which buffer fluids and/or dilution fluids may be retrieved forpresorting.

After the presorting stage, any desirable constituents 15 may bereturned along a return path 60 through the outlet 124 of the body-worndevice 120 to the biological fluid source 17 via the catheter 128. Anyundesirable constituents 14, or unhealthy cells, may be transferred tothe cartridge 126 along an isolation path 50. Any sample of biologicalfluid 16 after the presorting stage which includes a mixture ofrecognized fluid constituents 13 and undesirable constituents 14, suchas a mixture of healthy cells and unhealthy cells, are transferred tothe fluid receiving device 30 to be optically scanned by the scanner 70and processed by the control unit 80 in the manners described elsewhereherein.

FIG. 48 illustrates a method of filtering a biological fluid 16 within abody-worn device 120 which includes a drug infuser 130 in the returnpath 60. As shown in FIG. 48, a drug infuser 130 may be positioned alongthe return path 60 between the valve(s) 127 and the outlet 124 of thebody-worn device 120. Drugs or treatments may be infused by the druginfuser 130 with the filtered biological fluid 16 prior to returning tothe biological fluid source 17 through the outlet 124 of the body-worndevice 120 via the catheter 128. Various drugs or treatments may beutilized, and the body-worn device 120 may be configured such that thedrugs or treatments may be routinely re-filled or switched as-needed. Inother example embodiments, the body-worn device 120 may employadditional buffers or dilution fluids to presort the biological fluid16. Additionally, in some embodiments, the body-worn device 120 mayemploy anti-coagulants to prevent clotting.

FIG. 53 illustrates an exemplary method utilized for a multi-stagebody-worn filtration system in which the body-worn device 120 couldutilize optical filtration methods to filter and sequester samples ofbiological fluids 16 (e.g., blood) that contain CTCs and CTC-clusters.Such samples may be stored in a replaceable cartridge 126 that may thenbe further processed to enrich and extract CTCs and CTC-clusters.

As shown in FIG. 53, biological fluid 16 from a biological fluid source17 may be treated with optional anti-coagulant 49 prior to entering amicrofluidic separation module 100 for optional presorting. Anyundesirable constituents 14 are immediately transferred to an isolationpath 50. Any samples containing both undesirable constituents 14 anddesirable constituents 15 are transferred to a fluid receiving device 30and scanned by a scanner 70. The resulting scanned data 90 is processedby the control unit 80. If any undesirable constituents 14 are detected,the sample may be transferred along an isolation path 50 to be stored ina cartridge 126 for off-line enrichment to extract CTCs. If onlydesirable constituents 15 are detected, the sample may be returned tothe biological fluid source 17 by a return path 60.

FIG. 44C illustrates an exemplary portable device 140 which is portable(as opposed to body-worn) and may be utilized in a non-aphaereticsetting (e.g., to process cerebrospinal fluid, saliva, or urine toextract unhealthy cells such as CTCs). As shown in FIG. 44C, theportable device 140 will generally include a biological fluid inlet 142through which biological fluid 16 may be introduced into the portabledevice 140. An optional buffer fluid inlet 143 may be utilized tointroduce various buffer or dilation fluids for presorting of thebiological fluid 16. The buffer fluid may be replaced or replenished asneeded. Such an embodiment may also include anti-coagulants if utilizedto process fluids such as blood.

Continuing to reference FIG. 44C, the portable device 140 may includeindicators 144 (e.g., audible or visual) which provide variousinformation about the operability of the portable device 140 (e.g.,on/off, flow blockage, etc.). Internally to the portable device 140, afluid receiving device 30 and scanner 70 are provided for scanning thebiological fluid 16 within the portable device 140. Although not shown,the portable device 140 will generally include an internal power source,though the portable device 140 may be externally powered (e.g., by awall socket) in some embodiments. The portable device 140 may includeits own internal control unit or may be communicatively connected to aremote control unit for processing scanned data 90.

As shown in FIG. 44C, the portable device 140 may include a waste fluidoutlet 145 through which waste fluids and other disposables may beremoved from the portable device 140. The portable device 140 will alsogenerally include a removable cartridge 146 which stores any filteredcells. The cartridge 146 may be emptied and replaced as-needed toprocess additional biological fluids 16.

J. Closed-Loop Filtration, Treatment, and Monitoring

FIGS. 49 and 50 illustrate an exemplary embodiment of utilizing abiological fluid filtration system 10 for closed-loop filtration,treatment, and monitoring of undesirable constituents 14 such as harmfulcells. In such an embodiment, undesirable constituents 14 such as CTCsor CTC-clusters are filtered using the optical filtration methods andsystems described herein, with a drug or treatment (e.g.,chemotherapeutic agents, combination of drugs, immunotherapy treatments,etc.) being introduced in the return path 60 to check for itstherapeutic effect.

In an example embodiment, the ratio of undesirable constituents 14within a certain volume of biological fluid 16 may be monitoredcontinuously or sporadically to determine any changespost-administration of a certain drug or treatment. A decreasing ratioof undesirable constituents 14 such as viable (living) CTCs within abiological fluid 16 during aphaeresis may be an indication that acertain drug or treatment is effective.

For example, various optical scanning methods described herein (e.g.,digital holographic microscopy) may be utilized to count undesirableconstituents 14 per given volume of processed biological fluid 16. Insome embodiments, staining or fluorescence may be utilized. As with theother systems and methods described herein, the closed-loop filtration,treatment, and monitoring may be utilized in connection with humans oranimals.

FIG. 49 illustrates an exemplary method of filtering, monitoring, andtreating harmful cells within a biological fluid 16. As with the othersystems and methods described herein, a pre-filtration session may beperformed to improve the accuracy of the control unit 80 in analyzingthe biological fluid 16 and identifying any biological fluidconstituents 13. Data from the pre-filtration session may be utilized inwhole or in part in development of the reference data 91 which is usedby the control unit 80 to analyze biological fluid 16 and identify anybiological fluid constituents 13 as discussed herein. The reference data91 could include images and/or data of biological fluid constituents 13from the same patient or from other individuals.

As shown in FIG. 49, biological fluid 16 is drawn from a biologicalfluid source 17. Presorting may be applied as discussed herein, with anyundesirable constituents 14 being sequestered along an isolation path 50immediately after presorting. The remaining sample of biological fluid16 is then optically scanned by the scanner 70 and the resulting dataundergoes analysis by the control unit 80 to identify any biologicalfluid constituents 13.

Any samples of biological fluid 16 containing anything other thandesirable constituents 15 are transferred along the isolation path 50 tobe sequestered along any undesirable constituents 14 from the presorting(if performed). All such undesirable constituents 14 are monitored suchas by counting the number of viable CTCs or other harmful cells pervolume of processed biological fluid 16.

FIG. 50 illustrates an exemplary method of filtering, monitoring, andtreating harmful cells within a biological fluid 16 which includespresorting of cells prior to optical scanning, diagnostic profiling, andex vivo drug testing of undesirable constituents 14 to identify anoptimal drug or treatment to be administered in the return path 60. Inan exemplary embodiment, a drug or treatment may be tested on filteredcells by measuring changes to their cellular properties using opticaltechniques (e.g., digital holographic microscopy).

As an example embodiment, prior to drug administration, a portion of thefiltered cells may be processed for genomic data which aids inidentifying suitable drug targets. Those drugs are tested, potentiallyat various concentrations, on the remaining portion of the filteredcells to determine which drug or treatment has the most intendedtherapeutic effect so as to validate the optimal drug or treatmentchoice. Subsequently, the optimal drug or treatment is then introducedin the return path 60.

With reference to FIG. 50, it can be seen that a biological fluid 16 isdrawn from a biological fluid source 17, presorted, scanned, andanalyzed as described herein. Contents which are sequestered along theisolation path 50, such as undesirable constituents 14, undergodiagnostics such as cell counting (e.g., of viable CTCs), profiling(e.g., histochemistry, genomic transcriptomic, etc.), and the like. Thediagnostics are utilized to determine potential drugs or treatments.

After determining potential drugs or treatments based on diagnostics ofundesirable constituents 14 sequestered after presorting and opticalfiltration, optional drug sensitivity and resistance assay may beperformed. Based upon the results of the assay, optimal drugs ortreatments, including dosing, may be selected. Such optimal drugs ortreatments are then introduced in the return path 60 to be returned tothe biological fluid source 17.

Such an embodiment may be utilized to process multiple biological fluids16 from a biological fluid source 17 at the same time. Different typesof filtered cells may be held separately so that their counts,diagnostic profiling, and drug testing may be conducted separately.Optical scanning could include single or multiple parallel fluidreceiving devices 30. In some embodiments, the foregoing methods may beperformed on a sample of biological fluid 16.

K. Methods of Biological Fluid Filtration

Generally, the methods of biological fluid filtration are directed tothe removal of undesirable constituents 14 which may comprise, forexample, disease-related biological or chemical entities, from abiological fluid 16 by optically inspecting the constituents 13 in thebiological fluid 16 and filtering those that are not recognized to beamong the desirable constituents 15 of the biological fluid 16. Themethods of biological fluid filtration may be applied in a therapeuticcontext, a diagnostic context, or both, as described herein.

Generally, the methods involve directing a biological fluid 16 to afluid receiving device 30, optically scanning the biological fluid 16within the fluid receiving device 30 by a scanner 70 to generate thescanned data 90 of the biological fluid 16, comparing the scanned data90 of the biological fluid 16 with the reference data 91 by the controlunit 80; returning the biological fluid 16 to the biological fluidsource 17 if the scanned data 90 of the biological fluid 16 includesonly desirable constituents 15 exhibiting criteria that sufficientlymatch with any of the desirable constituents 15 of the reference data 91by the control unit 80; and isolating the biological fluid 16 from thebiological fluid source 17 if the scanned data 90 of the biologicalfluid 16 includes one or more constituents 13 not exhibiting criteriathat sufficiently match with any of the desirable constituents 15 of thereference data 91 by the control unit 80. The biological fluidfiltration system 10 described herein may be used to carry out themethods.

In one embodiment, the method of biological fluid filtration includespre-processing of the biological fluid 16 to separate particularcellular constituents of the biological fluid 16 for promotion to thefluid receiving device 30, as described above. Methods may alsooptionally include additional filtration processes, including antigenbased filtration, described above.

In an alternative embodiment, the method includes pre-processing of afluid sample of the biological fluid source 17 to obtain the referencedata 91 differentiating cell image data characteristics of desirableconstituents 15 to more precisely tailor the reference data 91 thebiological fluid system, including any heterogeneity among its desirableconstituents 15. Such pre-processing may rely upon machine learningand/or artificial intelligence models in order to more accurately andefficiently differentiate between undesirable constituents 14 anddesirable constituents 15.

In an alternative embodiment, the reference data 91 is developed basedon relevant normative subpopulation data concerning the desirableconstituents 15 of the biological fluid system.

L. Non-Aphaeretic Filtration Methods

It should be appreciated that, while many of the embodiments describedherein utilize aphaeretic systems, additional embodiments may benon-aphaeretic. One such exemplary embodiment is shown in FIG. 42, inwhich a non-aphaeretic implementation is utilized for the removal oftumor cell contaminants from analogous stem-cell transplant products.Another exemplary embodiment is shown in FIG. 44C, in which a portabledevice 140 may be utilized for non-aphaeretic filtration.

As shown in FIG. 42, biological fluid 16 comprised of a leukapheresisproduct may be collected from a biological fluid source 17. Pre-sortingmay be performed by a microfluidic separation module 100 so as toseparate the leukapheresis product into three primary divisions: (1)separated red blood cells, plasma, and small cells; (2) separated whiteblood cells and small CTCs; and (3) separated large CTC-clusters. Insome embodiments, pre-sorting may involve the use of chemical agentsand/or buffer solutions for red blood cell and platelet separation orlysis.

The first division, comprised of separated red blood cells, plasma, andsmall cells, may be transferred to a fluid chamber to hold filteredplasma and healthy cells for transplantation. The third division,comprised of large CTC-clusters, may be transferred for diagnosticprocessing (e.g., genomic, transcriptomic, metabolomics, drugsensitivity and resistance) of CTCs, CTC-clusters, and cell-free plasma.

With respect to the second division, comprised of separated white bloodcells and small CTCs, such contents may be inertial focused into one ormore parallel fluid receiving devices 30 (e.g., microfluidic channels31, microwell arrays 32, and/or droplet generators 34). Such contentsmay be scanned simultaneously or sequentially by the scanner 70. Thecontrol unit 80 will then analyze each sample to determine if cellsother than healthy cells are present (e.g., undesirable constituents14).

If no cells other than healthy cells are present in a given sample, thatsample may be transferred to a fluid chamber along with the firstdivision of separated red blood cells, plasma, and small cells. If cellsother than healthy cells are present in a given sample, that sample maybe further enriched (e.g., by returning to be reprocessed by areprocessing path 62) or may be sequestered along with the thirddivision of separated large CTC-clusters for diagnostic processing.

M. Operation of Illustrative Embodiments

The systems and methods described herein may be utilized for therapeuticpurposes, diagnostic purposes, or both. It should be appreciated thatthe methods and systems used to isolate biological fluids 16 containingundesirable constituents 14 may vary in different embodiments. Further,the methods and systems utilized for processing the isolated biologicalfluids 16 containing undesirable constituents 14 may vary in differentembodiments depending upon whether such isolated biological fluids 16are to be used for therapeutic purposes, diagnostic purposes, or somecombination of both.

i. Therapeutic Systems and Methods.

As shown throughout the figures and discussed herein, the systems andmethods described herein may be utilized for various therapeuticpurposes. For example, therapeutic embodiments in which the undesirableconstituents 14 are CTCs or CTC-clusters may be useful to reducemetastatic disease. As another example, therapeutic embodiments in whichthe undesirable constituents 14 are pathogens may be useful to treatinfection, including sepsis.

ii. Aphaeretic Scanning and Filtration of Whole Blood Fluid with CTCs ona Microfluidic Channel Platform with Reference Image Dataset of SampledBlood from Representative Individuals Other than the Patient.

One illustrative embodiment is a system and method for aphaereticscanning and filtration of whole blood to remove CTCs from a patient'scirculatory system. As illustrated in FIGS. 12 and 27, whole blood ispumped from a patient 12 via a receiver path 20 to a fluid receivingdevice 30 comprising a batch of parallel microfluidic channels 31. Eachchannel 31 is optically scanned by a scanner 70 using DIC Microscopy,DHM or other appropriate imaging techniques to derive a scanned data 90of the cells for each microfluidic channel 31.

The scanned data 90 is transferred to a control unit 80, which utilizesan image processing software program comprising an algorithm 82 designedto recognize healthy blood cells (i.e., erythrocytes, leukocytes andplatelets) in the scanned data 90. More particularly, the algorithm 82is designed to recognize patterns in the reference data 91characteristic of each type of healthy blood cell and process eachscanned data 90 transferred to the control unit 80 to determine whetherthose patterns are recognized in discreet image data obtained for eachcell.

In an exemplary embodiment, the reference data 91 is obtained throughDIC Microscopy, digital holographic microscopy, or other appropriateimaging techniques of blood samples taken from a representative sampleof individuals other than the patient. Raw data is computer-processed toidentify patterns in the images and/or data that reflect characteristicsof each type of healthy blood cell, including, for example, cell size,shape, texture, phase deviations, solidity and luminance. Datareflecting those patterns is uploaded to the control unit 80 and storedto memory as the reference data 91.

With reference to FIG. 29, if all cells in a scanned microfluidicchamber are recognized by the algorithm 82 as healthy blood cells, thenthe image processing software program generates a first control signalinstruction 83 to the control unit 80 to relay a control signal to thevalve 40 to route channel contents to the return path 60. If, on theother hand, one or more of cells of the scanned microfluidic chamber 31are not recognized by the algorithm 82, then the image processingsoftware program generates a second control signal instruction 84 to thecontrol unit 80 to relay a control signal 81 to the valve 40 to routechannel contents to the isolation path 50. Filtered blood in the returnpath 60 is pumped back to the patient's circulatory system. CTC-richfluid routed to the isolation path 50 is sequestered and optionallystored for further processing for diagnostic or therapeutic purposes.

iii. Aphaeretic Filtration of Leukocyte-Rich Blood Fluid with Small CTCson a Microfluidic Channel Platform with Reference Image Dataset ofSampled Blood from Representative Individuals Other than the Patient.

One illustrative embodiment is a system and method for aphaereticscanning and filtration of Leukocyte-Rich Blood Fluid to remove CTCsfrom a patient's circulatory system. In this example, which isillustrated in FIGS. 21 and 27, whole blood is pumped from the patient12 into a receiver path 20 that directs flow of the whole blood to amicrofluidic separation module 100, which pre-sorts whole blood usingappropriate sorting techniques, e.g., Dean flow fractionation ordielectric sorting, into three components: (1) fluid containingprimarily healthy erythrocytes (RBCs) and platelets, (2) fluid largelycontaining a mixture of leukocytes (WBCs) and small CTCs, and (3) fluidcontaining large CTCs and CTC-clusters. The fluid containing a mixtureof leukocytes and CTCs are promoted to fluid receiving device 30comprising a batch of parallel microfluidic channels 31. Each channel isoptically scanned using DIC Microscopy, DHM, or other appropriateimaging techniques to derive a scanned data 90 of the cells for eachmicrofluidic channel 31.

The scanned data 90 is transferred to a control unit 80, which followsan image processing software program comprising an algorithm 82 designedto recognize healthy blood cells in the scanned data 90. Moreparticularly, the algorithm 82 is designed to recognize a pattern in areference data 91 characteristic of healthy blood cells and process eachscanned data 90 transferred to the control unit 80 to determine whetherthat pattern is recognized in discreet image data obtained for eachcell. In the example, the reference data 91 is obtained through DICMicroscopy or DHM of blood samples taken from a representative sample ofindividuals other than the patient.

With reference to FIG. 29, if all cells in a scanned microfluidicchannel 31 are recognized by the algorithm 82 as healthy blood cells,then the image processing software program generates a first controlsignal instruction 83 to the control unit 80 to relay a control signal81 to the valve 40 to route channel contents to the return path 60. If,on the other hand, one or more of cells of the scanned microfluidicchannel are not recognized by the algorithm 82, then the imageprocessing software program generates a second control signalinstruction 84 to the control unit 80 to relay a control signal to thevalve 40 to route channel contents to the isolation path 50. Leukocytesrouted to the return path 60 are then recombined with the pre-sortedRBCs, plasma and platelets and pumped back to the patient's circulatorysystem. CTC-rich fluid routed to the isolation path 50 is sequesteredand optionally stored for further processing for diagnostic ortherapeutic purposes.

iv. Aphaeretic Scanning and Filtration of Whole Blood Fluid with CTCs ona Microfluidic Channel Platform with Reference Image Data of SampledBlood from Patient.

One illustrative embodiment is a system and method or aphaereticscanning and filtration of whole blood to remove CTCs from a patient'scirculatory system. As illustrated in FIGS. 12 and 28, whole blood ispumped from a patient 12 via a receiver path 20 to a fluid receivingdevice 30 comprising a batch of parallel microfluidic channels 31. Eachchannel 31 is optically scanned by a scanner 70, such as by using DICMicroscopy, DHM, or other appropriate imaging techniques, to derive ascanned data 90 of the cells for each microfluidic channel 31.

The scanned data 90 is transferred to a control unit 80, which followsan image processing software program comprising an algorithm 82 designedto recognize healthy blood cells in the scanned data 90. In the example,the reference data 91 is obtained through DIC Microscopy, DHM, or otherappropriate imaging techniques of blood samples taken from the patient12 prior to the filtration session. The algorithm 82 is designed torecognize patient-specific patterns in the reference data 91characteristic of each type of healthy blood cell and process eachscanned data 90 transferred to the control unit 80 to determine whetherthose patterns are recognized in discrete image data obtained for eachcell.

With reference to FIG. 29, if all cells in a scanned microfluidicchannel 31 are recognized by the algorithm 82 as healthy blood cells,then the image processing software program generates a first controlsignal instruction 83 to the control unit 80 to relay a control signalto the valve 40 to route channel contents to the return path 60 If, onthe other hand, one or more of cells of the scanned microfluidic channelare not recognized by the algorithm 82, then the image processingsoftware program generates a second control signal instruction 84 to thecontrol unit 80 to relay a control signal to the valve 40 to routechannel contents to the isolation path 50. Filtered blood in the returnpath 60 is pumped back to the patient's circulatory system. CTC-richfluid routed to the isolation path 50 is sequestered and optionallystored for further processing for diagnostic or therapeutic purposes.

v. Aphaeretic Filtration of Leukocyte-Rich Blood Fluid with Small CTCson a Microfluidic Channel Platform with Reference Image Data of SampledBlood from Patient.

One illustrative embodiment is a system and method for aphaereticscanning and filtration of leukocyte-rich blood fluid to remove CTCsfrom a patient's circulatory system. In this example, as illustrated inFIGS. 17 and 28, whole blood is pumped from the patient into a receiverpath 20 that directs flow of the whole blood to a microfluidicseparation module 100, which pre-sorts whole blood using appropriatesorting techniques, e.g., Dean flow fractionation or dielectric sorting,into three components: (1) fluid containing primarily healthyerythrocytes (RBCs) and platelets, (2) fluid largely containing amixture of leukocytes (WBCs) and small CTCs, and (3) fluid containinglarge CTCs and CTC-clusters. The fluid containing a mixture ofleukocytes and CTCs are promoted to fluid receiving device 30 comprisinga batch of parallel microfluidic channels 31. Each channel is opticallyscanned by a scanner 70 which may, for example, use DIC Microscopy, DHM,or other appropriate imaging techniques to derive a scanned data 90 ofthe cells for each microfluidic channel 31.

The scanned data 90 is transferred to a control unit 80, which followsan image processing software program comprising an algorithm 82 designedto recognize healthy blood cells in the scanned data 90. In the example,a reference data 91 is obtained through DIC Microscopy, DHM, or otherappropriate imaging techniques of blood samples taken from the patientprior to the filtration session. The algorithm 82 is designed torecognize a pattern in the reference data 91 characteristic of thepatient's healthy cells and process each scanned data 90 transferred tothe control unit 80 to determine whether that pattern is recognized indiscreet image data obtained for each cell.

With reference to FIG. 29, if all cells in a scanned microfluidicchannel 31 are recognized by the algorithm 82 as healthy cells, then theimage processing software program generates a first control signalinstruction 83 to the control unit 80 to relay a control signal 81 tothe valve 40 to route channel contents to the return path 60. If, on theother hand, one or more of cells of the scanned microfluidic channel arenot recognized by the algorithm 82, then the image processing softwareprogram generates a second control signal instruction 84 to the controlunit 80 to relay a control signal 81 to the valve 40 to route channelcontents to the isolation path 50. Leukocytes routed to the return path60 are then recombined with the pre-sorted RBCs, plasma and plateletsand pumped back to the patient's circulatory system. CTC-rich fluidrouted to the isolation path 50 is sequestered and optionally stored forfurther processing for diagnostic or therapeutic purposes.

vi. Aphaeretic Scanning and Filtration of Whole Blood Fluid withPathogens on a Microfluidic Channel Platform.

An illustrative embodiment is a system and method for aphaereticscanning and filtration of whole blood to remove pathogens from apatient's circulatory system. As shown in FIGS. 15 and 27-28, wholeblood is pumped from a patient 12 via a receiver path 20 to a fluidreceiving device 30 comprising a batch of parallel microfluidic channels31. Each channel is optically scanned using DIC Microscopy, DHM, orother appropriate imaging techniques to derive a scanned data 90 of thecells for each microfluidic channel 31.

The scanned data is transferred to a control unit 80, which utilizes animage processing software program comprising an algorithm 82 designed torecognize healthy blood cells in the scanned data 90. More particularly,the algorithm 82 is designed to recognize patterns in reference imagedata of the reference data 91 characteristic of each type of healthyblood cell and process each scanned data 90 transferred to the controlunit 80 to determine whether those patterns are recognized in discreetimage data obtained for each cell. The reference data 91 is obtainedthrough DIC Microscopy, DHM, or other appropriate imaging techniques ofeither samples taken from the patient's blood or samples taken from arepresentative sample of individuals other than the patient.

With reference to FIG. 29, if all cells in a scanned microfluidicchannel 31 are recognized by the algorithm 82 as healthy blood cells,then the image processing software program generates a first controlsignal instruction 83 to the control unit 80 to relay a control signal81 to the valve 40 to route channel contents to the return path 60. If,on the other hand, one or more of cells of the scanned microfluidicchannel 31 are not recognized by the algorithm, then the imageprocessing software program generates a second control signalinstruction 84 to the control unit 80 to relay a control signal 81 tothe valve 40 to relay a control signal to the valve 40 to route channelcontents to the isolation path 50. Filtered blood in the return path 60is pumped back to the patient's circulatory system. Pathogen-infectedfluid in the isolation path 50 is sequestered and optionally stored forfurther processing for diagnostic or therapeutic purposes

vii. Aphaeretic Filtration of Pre-Sorted Blood Fluid with Pathogens onMicrofluidic Channel Platform.

An illustrative embodiment is a system and method for aphaereticfiltration of Pre-Sorted Blood Fluid to remove pathogens from apatient's circulatory system. In this example, as shown in FIGS. 15 and27-28, whole blood is pumped from the patient 12 into a receiver path 20that directs flow of the whole blood to a microfluidic separation module100, which pre-sorts whole blood using appropriate sorting techniques,e.g., Dean flow fractionation or dielectric sorting, into threecomponents: (1) fluid containing only healthy blood cells, (2) fluidcontaining a mixture of healthy blood cells and pathogens, and (3) fluidcontaining only pathogens. The fluid containing a mixture of blood cellsand pathogens are promoted to a fluid receiving device 30 comprising abatch of parallel microfluidic channels 31. Each channel is opticallyscanned using by a scanner 70 which uses DIC Microscopy, DHM, or otherappropriate imaging techniques to derive a scanned data 90 of cells foreach channel.

The scanned data 90 is transferred to a control unit 80, which utilizesan image processing software program comprising an algorithm 82 designedto recognize healthy blood cells in the scanned data 90. Moreparticularly, the algorithm 82 is designed to recognize a patterns in areference data 91 characteristic of each type of healthy blood cell andprocess each scanned data 90 transferred to the control unit todetermine whether that pattern is recognized in discreet image dataobtained for each cell. The scanned data 90 may be obtained through DICMicroscopy, DHM, or other appropriate imaging techniques of eithersamples taken from the patient's blood or samples taken from arepresentative sample of individuals other than the patient.

With reference to FIG. 29, if all cells in a scanned microfluidicchannel 31 are recognized by the algorithm as healthy blood cells, thenthe image processing software program generates a first control signalinstruction 83 to the control unit 80 to relay a control signal 81 to avalve 40 to route the channel's contents to the return path 60. If, onthe other hand, one or more of cells of the scanned channel 31 are notrecognized by the algorithm 82, then the image processing softwareprogram generates a second control signal instruction 84 to the controlunit 80 to relay a control signal 81 to the valve 40 to route thechannel's contents to the isolation path 50. Blood cells routed to thereturn path 60 are then recombined with the pre-sorted healthy bloodcells and pumped back to the patient's circulatory system.Pathogen-infected fluid routed to the isolation path 50, along with thepre-sorted fluid containing only pathogens, are sequestered andoptionally stored for further processing for diagnostic or therapeuticpurposes.

viii. Aphaeretic Scanning and Filtration of Whole Blood Fluid with CTCson a Microwell Array.

An illustrative embodiment is a system and method for aphaereticscanning and filtration of whole blood to remove CTCs from a patient'scirculatory system using a microwell array 32. As shown in FIGS. 8-11,and 27-28, whole blood is pumped from a patient 12 via a receiver path20 to a fluid receiving device 30 comprising a microwell array 32. Eachmicrowell of the microwell array 32 is optically scanned with a scanner70 using DIC Microscopy, DHM, or other appropriate imaging techniques toderive a scanned data 90 of the cells for each microwell of themicrowell array 32. Micro-wells may be periodically disrupted, such asby shaking or stirring, and multiple scans taken, to ensure that thescanned data 90 captures the entirety of the contents of the microwells.

The scanned data 90 is transferred to a control unit 80, which utilizesan image processing software program comprising an algorithm 82 designedto recognize healthy blood cells in the scanned data 90. Moreparticularly, the algorithm is designed to recognize patterns inreference image data of the reference data 91 characteristic of eachtype of healthy blood cell and process each scanned data 90 transferredto the control unit 80 to determine whether those patterns arerecognized in discreet image data from the reference data 91 obtainedfor each cell. The reference data 91 is obtained through DIC Microscopy,DHM, or other appropriate imaging techniques of blood samples taken fromeither from a sample of blood from the patient 12 or a representativesample of individuals other than the patient 12.

With reference to FIG. 29, if all cells in a scanned microwell of themicrowell array 32 are recognized by the algorithm as healthy bloodcells, then the image processing software program generates a firstcontrol signal instruction 83 to the control unit 80 to relay a controlsignal 81 to the valve 40 at the base of the microwell to route thecontents of the microwell to the return path 60 If, on the other hand,one or more of cells of the scanned microwell are not recognized by thealgorithm, then the image processing software program generates a secondcontrol signal instruction 84 to the control unit 80 to relay a controlsignal 81 to the valve 40 to route the contents of the microwell to theisolation path 50. Filtered blood in the return path 60 is pumped backto the patient's circulatory system. CTC-rich fluid routed to theisolation path 50 is sequestered and optionally stored for furtherprocessing for diagnostic or therapeutic purposes. In some exampleembodiments, pipettes could be utilized to extract contents of amicrowell. In other embodiments, multiple wells in an array could sharea single valve 40.

ix. Aphaeretic Filtration of Leukocyte-Rich Blood Fluid with Small CTCson a Microwell Array.

An illustrative embodiment is a system and method for aphaereticscanning and filtration of leukocyte-rich blood fluid to remove CTCsfrom a patient's circulatory system using a microwell array 32. Withreference to FIGS. 8-10 and 27-28, whole blood is pumped from thepatient 12 into a receiver path 20 that directs flow of the whole bloodto a microfluidic separation module 100, which pre-sorts whole bloodusing appropriate sorting techniques, e.g., Dean flow fractionation ordielectric sorting, into three components: (1) fluid containing onlyhealthy erythrocytes (RBCs) and platelets, (2) fluid largely containinga mixture of leukocytes (WBCs) and small CTCs, and (3) fluid containinglarge CTCs and CTC-clusters. The fluid containing a mixture ofleukocytes and CTCs are promoted to a fluid receiving device 30comprising a microwell array 32. Each microwell is optically scanned bya scanner 70, which may use DIC Microscopy, DHM, or other appropriateimaging techniques to derive a scanned data 90 of the cells for eachmicrowell.

The scanned data 90 is transferred to a control unit 80, which utilizesan image processing software program comprising an algorithm 82 designedto recognize healthy blood cells in the scanned data 90. Moreparticularly, the algorithm 82 is designed to recognize a pattern in areference data 91 characteristic of healthy blood cells and process eachscanned data 90 transferred to the control unit 80 to determine whetherthat pattern is recognized in discreet image data obtained for eachcell. The reference data 91 is obtained through DIC Microscopy of asample of the patient's blood or blood samples taken from arepresentative sample of individuals other than the patient 12.

With reference to FIG. 29, if all cells in a scanned microwell arerecognized by the algorithm 82 as healthy blood cells, then the imageprocessing software program generates a first control signal instruction83 to the control unit 80 to relay a control signal 81 to the valve 40to route the contents of the microwell to the return path 60. If, on theother hand, one or more of cells of the scanned microwell are notrecognized by the algorithm 82, then the image processing softwareprogram generates a second control signal instruction 84 to the controlunit 80 to relay a control signal 81 to the valve 40 to route thecontents of the microwell to isolation path 50. Leukocytes routed to thereturn path 60 are then recombined with the pre-sorted RBCs, plasma andplatelets and pumped back to the patient's circulatory system. CTC-richfluid routed to the isolation path 50 is sequestered and optionallystored for further processing for diagnostic or therapeutic purposes. Insome example embodiments, pipettes could be utilized to extract contentsof a microwell. In other embodiments, multiple wells in an array couldshare a single valve 40.

x. Aphaeretic Filtration of Leukocyte-Rich Blood Fluid with Small CTCson a Microfluidic Channel Platform with Reference Image Data for CTCs.

An illustrative embodiment is a system and method for aphaereticscanning and filtration of Leukocyte-Rich Blood Fluid to remove CTCsfrom a patient's circulatory system using reference data 91 of healthyblood cells and CTCs. With reference to FIG. 20, a biological fluid 16comprised of whole blood is pumped from the patient 12 into a receiverpath 20 that directs flow of the whole blood to a microfluidicseparation module 100, which pre-sorts whole blood using appropriatesorting techniques, e.g., Dean flow fractionation or dielectric sorting,into three components: (1) fluid containing only healthy erythrocytes(RBCs) and platelets, (2) fluid largely containing a mixture ofleukocytes (WBCs) and small CTCs, and (3) fluid containing large CTCsand CTC-clusters. The fluid 16 containing a mixture of leukocytes andCTCs are promoted to a fluid receiving device 30 comprising a batch ofparallel microfluidic channels 31. Each channel 31 is optically scannedwith a scanner 70 using DIC Microscopy, DHM, or other appropriateimaging techniques to derive a scanned data 90 of the cells for eachmicrofluidic channel 31.

The scanned data 90 is transferred to a control unit 80, which utilizesan image processing software program comprising an algorithm 82 designedto recognize healthy blood cells and CTCs in the scanned data 90. Moreparticularly, the algorithm 82 is designed to recognize a pattern in areference data 91 characteristic of healthy blood cells and CTCs andprocess each scanned data 90 transferred to the control unit 80 todetermine whether that pattern is recognized in discreet image dataobtained for each cell. The reference data 91, including image data ofrecognized CTCs, is obtained through DIC Microscopy, DHM, or othermethods of blood samples containing CTCs taken from either from a sampleof blood from the patient or a representative sample of individualsother than the patient. Image data from the reference data 91 may alsoinclude scanned images of CTCs appropriately cultured in medium or otherfluids.

With reference to FIG. 29, if all cells in a scanned microwell arerecognized by the algorithm 82 as all healthy blood cells, then theimage processing software program generates a first control signalinstruction 83 to the control unit 80 to relay a control signal 81 tothe valve 40 to route the contents of the channel to the return path 60.If, on the other hand, one or more of cells of the scanned microwell arerecognized as CTCs or not recognized by the algorithm 82, then the imageprocessing software program generates a second control signalinstruction 84 to the control unit 80 to relay a control signal 81 tovalve 40 to route the contents of the channel to isolation path 50.Leukocytes routed to the return path 60 are then recombined with thepre-sorted RBCs, plasma and platelets and pumped back to the patient'scirculatory system. CTC-rich fluid routed to the isolation path 50 issequestered and optionally stored for further processing for diagnosticor therapeutic purposes.

xi. Aphaeretic Scanning and Filtration of Whole Blood Fluid with CTCs ona Microfluidic Channel Platform with Reference Image Data for CTCs forValidation.

An illustrative embodiment is a system and method for aphaereticscanning and filtration of whole blood to remove CTCs from a patient's12 circulatory system using reference image data from a reference data91 for both healthy blood cells and CTCs. With reference to FIG. 20,whole blood is pumped from a patient 12 via a receiver path 20 to afluid receiving device 30 comprising a batch of parallel microfluidicchannels 31. Each channel is optically scanned by a scanner 70 using DICMicroscopy, DHM, or other appropriate imaging techniques to derive ascanned data 90 of the cells for each microfluidic channel 31.

The scanned data 90 is transferred to a control unit 80, which utilizesan image processing software program comprising an algorithm 82 designedto recognize healthy blood cells in the scanned data 90 and a certifyingalgorithm 85 designed to recognize CTCs in the scanned data 90. In theexample, a reference data 91 is obtained through DIC Microscopy, DHM, orother appropriate imaging techniques of a sample of blood samplescontaining CTCs taken from either from a sample of blood from thepatient 12 or a representative sample of individuals other than thepatient 12. Image data of the reference data 91 may also include scannedimages of CTCs appropriately cultured in medium or other fluids.

With reference to FIG. 29, the algorithm 82 is designed to recognizepatterns in the reference data 91 characteristic of CTCs and each typeof healthy blood cell and process each scanned data 90 transferred tothe control unit 80 to determine whether those patterns are recognizedin discreet image data obtained for each cell and generate one of twocontrol signal instruction to the control unit: a first control signalinstruction 83 routing the channel's contents to the return path 60 ifevery cell in the channel is recognized as a healthy blood cell, and asecond control instruction 84 routing the channel's contends determinedto contain CTCs to the isolation path 50 if one or more cells in thechannel is not recognized as a healthy blood cell.

The certifying algorithm 85 is designed to generate a certificationoutput before a first control instruction 83 can be executed by thecontrol unit 80. The certifying algorithm 85 is designed to recognize apattern in the reference data 91 characteristic of the CTCs and verifythat the CTC pattern is not recognized in the scanned data 90 on which agenerated first control signal instruction 83 is based. If theverification condition is met, the certifying algorithm 85 generates acertification output and the control unit 80 is given a go instructionto execute the first control signal instruction 83. If the verificationcondition is not met, the certifying algorithm 85 generates an erroroutput, prompting the image processing software program to replace thefirst control signal instruction 83 with a second control signalinstruction 84 executed by the control unit 80. Filtered blood in returnpath is pumped back to the patient's circulatory system. CTC-rich fluidrouted to the isolation path 50 is sequestered and may optionally befurther processed for diagnostic or therapeutic purposes.

In some embodiments, the biological fluid 16 within the fluid receivingdevice 30 may be scanned multiple times to verify that the biologicalfluid 16 within the fluid receiving device 30 is indeed healthy prior toreturning to the biological fluid source 17 via the return path 60. Insuch an embodiment, after an initial scan of the biological fluid 16 bythe scanner 70 fails to detect any undesirable constituents 14, thebiological fluid 16 will be re-scanned, either by the same scanner 70 orby a different scanner 70 in embodiments with multiple scanners 70, toverify and confirm the absence of any undesirable constituents 14 withinthe biological fluid 16. The number of times that the biological fluid16 is scanned in such a verification algorithm may vary in differentembodiments.

xii. Aphaeretic Filtration of Leukocyte-Rich Blood Fluid with Small CTCson a Microfluidic Channel Platform with Reference Image Data for CTCsfor Validation.

An illustrative embodiment is a system and method for aphaereticscanning and filtration of Leukocyte-Rich Blood Fluid to remove CTCsfrom a patient's circulatory system using reference image data from areference data 91 of healthy blood cells and CTCs. With reference toFIGS. 20 and 27-28, biological fluid 16 comprised of whole blood ispumped from the patient 12 into a receiver path 20 that directs flow ofthe whole blood to a microfluidic separation module 100, which pre-sortswhole blood using appropriate sorting techniques, e.g., Dean flowfractionation or dielectric sorting, into three components: (1) fluidcontaining primarily healthy erythrocytes (RBCs) and platelets, (2)fluid largely containing a mixture of leukocytes (WBCs) and small CTCs,and (3) fluid containing large CTCs and CTC-clusters. The fluidcontaining a mixture of leukocytes and CTCs are promoted to a fluidreceiving device 30 comprising a batch of parallel microfluidic channels31. Each channel is optically scanned by a scanner 70 using DICMicroscopy, DHM, or other appropriate imaging techniques to derive ascanned data 90 of the cells for each microfluidic channel 31.

The scanned data 90 is transferred to a control unit 80, which utilizesan image processing software program comprising an algorithm 82 designedto recognize healthy blood cells in the scanned data 90 and a certifyingalgorithm 85 designed to recognize CTCs in the scanned data 90. In theexample, a reference data 91 is obtained through DIC Microscopy, DHM, orother appropriate imaging techniques of blood samples containing CTCstaken from either from a sample of blood from the patient 12 or arepresentative sample of individuals other than the patient 12. Thereference data 91 may also include scanned images of CTCs appropriatelycultured in medium or other fluids.

With reference to FIG. 29, the algorithm 82 is designed to recognizepatterns in the reference data 91 characteristic of healthy blood cellsand process each scanned data 90 transferred to the control unit 80 todetermine whether those patterns are recognized in discreet image dataobtained for each cell and generate one of two control signalinstruction to the control unit: a first control signal instruction 83to route channel contents to the return path 60 if every cell in thechannel is recognized as healthy blood cell, and a second controlinstruction 84 to route channel contends determined to contain CTCs tothe isolation path 50 if one or more cells in the channel is notrecognized as a healthy blood cell. The certifying algorithm 85 isdesigned to generate a certification output before a first controlinstruction can be executed by the control unit 80.

The certifying algorithm 85 is designed to recognize a pattern in thereference data 91 characteristic of CTCs and verify that the CTC patternis not recognized in a scanned data 90 on which a generated firstcontrol signal instruction 83 is based. If the verification condition ismet, the certifying algorithm 85 generates a certification output andthe control unit 80 is given a go instruction to execute the firstcontrol signal instruction 83. If the verification condition is not met,the certifying algorithm generates an error output, prompting the imageprocessing software program to replace the first control signalinstruction 83 with a second control signal instruction 84 executed bythe control unit 80. Filtered blood in return path is pumped back to thepatient's circulatory system. CTC-rich fluid routed to the isolationpath 50 is sequestered and may optionally be further processed fordiagnostic or therapeutic purposes.

In some embodiments, the biological fluid 16 within the fluid receivingdevice 30 may be scanned multiple times to verify that the biologicalfluid 16 within the fluid receiving device 30 is indeed healthy prior toreturning to the biological fluid source 17 via the return path 60. Insuch an embodiment, after an initial scan of the biological fluid 16 bythe scanner 70 fails to detect any undesirable constituents 14, thebiological fluid 16 will be re-scanned, either by the same scanner 70 orby a different scanner 70 in embodiments with multiple scanners 70, toverify and confirm the absence of any undesirable constituents 14 withinthe biological fluid 16. The number of times that the biological fluid16 is scanned in such a verification algorithm may vary in differentembodiments.

xiii. Aphaeretic Scanning and Filtration of Blood with CTCs on aMicrofluidic Channel Platform Including Reference Image Data to LimitFalse Positives.

An illustrative embodiment is a system and method for aphaereticscanning and filtration of blood fluid to remove undesirable CTCs orpathogens from a patient's circulatory system using reference image datathat includes recognized patterns for optic artifacts and designatedpathogens not intended for filtration to limit erroneous second controlsignal instructions 84 based on false positives.

In the example, as shown in FIGS. 16 and 27-28, reference data 91 forhealthy blood cells is obtained through DIC Microscopy, DHM, or otherappropriate imaging techniques, as described herein. The reference data91 also includes image data of known optic artifacts and scanned data 90of known non-target blood borne pathogens. The image processing softwareprogram of the control unit 80 of this embodiment comprises an algorithm82 designed to recognize patterns in the reference data 91characteristic of each type of healthy blood cell and to recognize datapatterns of optic artifacts and designated pathogens in reference data91 to avoid interpreting these data patterns as undesirable CTCs.

In operation, blood fluid on the fluid receiving device 30 is opticallyscanned, as described herein, to obtain a scanned data 90 of the cellsin the scanned biological fluid 16. The scanned data 90 is transferredto the control unit 80, and the algorithm 82 processes the scanned data90 to determine whether patterns for healthy blood cells, opticartifacts, and designated pathogens in the reference data 91 isrecognized in discreet image data of the scanned data 90 for each cell.As with other embodiments, the image processing software programgenerates a first control signal instruction 83 if all cells arerecognized by the algorithm 82, or a second control signal instruction84 if one or more cells is not recognized. Because recognized patterndata of optic artifacts and designated pathogens is incorporated intothe reference data 91, the presence of optic artifacts in the scanneddata 90 or of non-target pathogens in the scanned biological fluid willnot cause algorithm 82 to trigger the second control signal instruction84.

It is preferable to maximize the amount of healthy blood cells returnedto the patient 12 and, relatedly, to limit the amount of healthy bloodcells lost during filtration. The biological fluid filtration system 10operates most efficiently when scanned cells routed to the return pathbased on a first control signal instruction 83 are all healthy bloodcells and when scanned cells routed to the isolation path 50 based on afirst control signal instruction 83 comprise at least one undesirableCTC that triggered the instruction. Image data from optical artifacts orblood borne pathogens are often picked up by optic scans, and so havethe potential to trigger a first control signal instruction 83. Thepresent embodiment mitigates this issue by adding image data fromoptical artifacts or designated pathogens to the reference data 91recognized algorithm 82.

xiv. Diagnostic System and Methods.

As shown throughout the figures and discussed herein, the systems andmethods described herein may be utilized for various diagnosticpurposes. For example, undesirable constituents 14 may be sequesteredand subject to a variety of in vitro diagnostic tools for purposes ofidentification, prognosis, and/or treatment determinations, within anumber of methodological categories, including microscopy,immunology-based assaying, culturing, in vitro testing, drug sensitivityand resistance testing, and genomic testing.

xv. Scanning and Filtration of Blood Sample for Diagnosis of aPathogenic Infection.

FIG. 14 is a block diagram of a diagnostic system and method of anembodiment adapted to remove undesirable, indeterminate pathogens from abiological fluid 16 and processing the removed pathogens fordiagnostics.

In the example, a blood sample is loaded into the fluid receiving device30, which comprises a batch of parallel microfluidic channels 31 adaptedto receive the blood sample. Each channel 31 is optically scanned by ascanner 70 using DIC Microscopy, DHM, or other appropriate imagingtechniques to derive a scanned data 90 of the cells for eachmicrofluidic channel 31.

The scanned data 90 is transferred to a control unit 80 which follows animage processing software program comprising an algorithm 82 designed torecognize healthy blood cells in the scanned data 90. In the example,reference data 91 is obtained through DIC Microscopy, DHM, or otherappropriate imaging techniques of blood samples taken from the patient12 or from a representative sample of individuals other than the patient12 prior to the diagnostic session. The algorithm 82 is designed torecognize a pattern in the reference data 91 and process each scanneddata 90 transferred to the control unit 80 to determine whether thatpattern is recognized in discreet image data from the scanned data 90obtained for each cell.

If all cells in a scanned microfluidic channel 31 are recognized by thealgorithm 82 as healthy blood cells, the control unit 80 causes thevalve 40 to direct contents of the channel 31 for disposal. If, on theother one or more of cells of the scanned microfluidic channel 31 arenot recognized by the algorithm 82, the control unit 80 causes the valve40 to direct the fluid 16 for collection and diagnostic processing.

The control unit 80 is also adapted collect and store catalogued imagedata for a group of catalogued pathogens, and the algorithm 82 isfurther adapted to recognize patterns in the reference data 91characteristic of catalogued pathogens, and process a scanned data 90 tosearch those patterns in the scanned data 90, and identify anycatalogued pathogen whose pattern is recognized in the scanned data 90.The control unit 80 is further adapted to store scanned data 90 to thecatalogued image data, assign the scan image data to a cataloguedpathogen whose pattern is recognized in the scan image data or to apathogen later identified in further diagnostic procedures, and thealgorithm 82 is further adapted to recognize the scanned data 90 for usein connection in with diagnostic procedures.

xvi. Scanning and Filtration of Blood Sample for Diagnosis of a Cancer.

FIG. 14 also illustrates an embodiment diagnostic system and methodadapted to remove undesirable indeterminate CTCs or CTC-clusters from abiological fluid 16 and process the removed CTCs or CTC-clusters fordiagnostics. Such an embodiment could be utilized to detect the presenceof cancer in an individual.

In the example, a blood sample is loaded into the fluid receiving device30, which comprises a batch of parallel microfluidic channels 31 adaptedto receive the blood sample. Each channel 31 is optically scanned by ascanner 70 using DIC Microscopy, DHM, or other appropriate imagingtechniques to derive a scanned data 90 of the cells for eachmicrofluidic channel 31.

The scanned data 90 is transferred to a control unit 80 which utilizesan image processing software program comprising an algorithm 82 designedto recognize healthy blood cells in the scanned data 90. In the example,a reference data 91 is obtained through DIC Microscopy, DHM, or otherappropriate imaging techniques of blood samples taken from the patient12 or from a representative sample of individuals other than the patient12 prior to the diagnostic session. The algorithm 82 is designed torecognize a pattern in the reference data 91 characteristic of healthyblood cells and process each scanned data 90 transferred to the controlunit 80 to determine whether that pattern is recognized in discreetimage data from the scanned data 90 obtained from each cell.

If all cells in a scanned microfluidic channel 31 are recognized by thealgorithm 82 as healthy blood cells, the control unit 80 causes thevalve 40 to direct contents of the channel 31 for disposal. If, on theother hand, one or more of cells of the scanned microfluidic channel 31are not recognized by the algorithm 82, the control unit 80 causes thevalve 40 to direct the fluid 16 for collection and diagnosticprocessing.

The control unit 80 is also adapted collect and store catalogued imagedata for a group of catalogued CTCs, and the algorithm 82 is furtheradapted to recognize patterns in the reference data 91 characteristic ofcatalogued CTCs, and process a scanned data 90 to search those patternsin the scanned data 90, and identify any catalogued CTCs whose patternis recognized in the scanned data 90. The control unit 80 is furtheradapted to store scan image data to the scanned data 90, assign thescanned data 90 to a catalogued CTC whose pattern is recognized in thescanned data 90 or to a CTC identified in further diagnostic procedures,e.g., PCR, and the algorithm 82 is further adapted to recognize thescanned data 90 for use in connection with diagnostic procedures.

xvii. Scanning and Filtration of Leukapheresis Extract.

FIG. 32 illustrates an exemplary embodiment of scanning and filtering aleukapheresis extract from a patient 12 undergoing leukapheresis. Asshown in FIG. 32, biological fluid 16 comprised of a leukapheresisextract from the patient 12 containing healthy cells and tumor cells isextracted from the patient 12. The leukapheresis extract may bepresorted using a microfluidic separation module 100 in which healthycells (e.g., utilizing RBC lysis, microfluidic sorting, etc.) areseparated from the leukapheresis extract.

The remaining sample of the leukapheresis extract after presorting isdirected into a fluid receiving device 30 (e.g., microfluidic channels31, microwell arrays 32, and/or droplet generators 34) and then scannedby the scanner 70. The scanned data 90 from the scanner 70 istransferred to the control unit 80 which, utilizing a reference data 91,detects the contents of the leukapheresis extract.

If cells other than healthy cells are detected within the scannedleukapheresis extract sample, the sample may be optionally returned by areprocessing path 62 for further enrichment of non-healthy cells. Suchsamples may also be processed for diagnostics. If only healthy cells aredetected within the scanned leukapheresis extract sample, the sample maybe directed along an isolation path 50 and not be processed fordiagnostics.

FIG. 37 illustrates another exemplary method of scanning and filtering aleukapheresis extract. As shown in FIG. 37, the presorting stagecomprised of a microfluidic separation module 100 may be utilized foroptional red blood cell and platelet separation using microfluidic orbiochemical methods. After scanning by the scanner 70 on the fluidreceiving device 30, any samples containing unrecognized cells may beprocessed for extraction of such unrecognized cells (e.g., T-cellextraction) and may optionally be reprocessed by redirecting back to thefluid receiving device 30 via a reprocessing path 62. Any samplescontaining only recognized healthy cells may be directed along anisolation path 50 not to be processed for cell extraction.

xviii. Scanning and Filtration of Stem Cell Extract.

FIG. 33 illustrates an exemplary embodiment of scanning and filtering astem cell extract from a patient 12 undergoing procedures for removal ofstem cells. As shown in FIG. 33, biological fluid 16 comprised of a stemcell extract is extracted from the patient 12 containing healthy cellsand any tumor cells. The stem cell extract may optionally be directed toa microfluidic separation module 100 for separation of red blood cellsand platelets using microfluidic or biochemical methods.

The remaining stem cell extract after the optional presorting stage isdirected to the fluid receiving device 30 in which it is opticallyscanned by a scanner 70. The scanned data 90 from the scanner 70 istransferred to the control unit 80 which, utilizing a reference data 91,detects the contents of the stem cell extract.

If cells other than healthy cells (e.g., tumor cells, pathogens, etc.)are detected in the stem cell extract sample, the sample may beoptionally directed along a reprocessing path 62 for optionalreprocessing for further enrichment of non-healthy cells. Any such stemcell extract samples (whether further enriched or not) may then beprocessed for diagnostics. Stem cell extract samples containing onlyhealthy cells (e.g., not including tumor cells or pathogens) may bedirected along an isolation path 50 to be utilized for stem celltransplant.

xix. Scanning and Filtration of Dialyzed Blood.

FIG. 51 illustrates an exemplary embodiment of scanning and filteringdialyzed blood from a dialysis unit 48. In such an embodiment, theoptical filtration unit is connected to a dialysis unit 48 (i.e., adialysis machine), where blood (either pre- or post-dialysis) is sent tothe optical filtration unit and healthy blood is returned back to thedialysis unit 48.

As shown in FIG. 51, biological fluid 16 from the biological fluidsource 17 is first undergoes dialysis within a dialysis unit 48. Theresulting biological fluid 16 which has undergone dialysis (e.g.,dialyzed blood) is then transferred to a fluid receiving device 30 andoptically scanned by a scanner 70. The resulting scanned data 90 isprocessed by the control unit 80. If only desirable constituents 15 aredetected, the sample may be returned back to the dialysis unit 48 by areturn path 60. If undesirable constituents 14 are detected, the samplemay be isolated along an isolation path 50 and are not returned to thedialysis unit 48. Optionally, diagnostics may be performed on thecontents (e.g., pathogens, CTCs, CTC-clusters, host cells, cell freeplasma, etc.) that have been isolated.

xx. Scanning and Filtration of Blood Bag Contents.

FIG. 52 illustrates an exemplary embodiment of scanning a biologicalfluid 16 comprised of blood from a biological fluid source 17 comprisedof a blood bag. In such an embodiment, the contents of a blood bag froma patient 12 are processed to filter CTCs and CTC-clusters. Theremaining contents may then be separated using a variety of methods(e.g., microfluidic, centrifugation, biochemical, etc.) to separatewhite blood cells and CTCs. Those cells may then be processed viaoptofluidic filtration techniques described herein to separate CTCs.

As shown in FIG. 52, a biological fluid source 17 comprised of a bloodbag may be utilized. The biological fluids 16 within the blood bag maybe transferred to a microfluidic separation module 100 for presorting oflarge CTC-clusters. Any such separated large CTC-clusters may betransferred along an isolation path 50 for diagnostics (e.g., count,viability, genomic, transcriptomic, molecular, morphological analyses).

The microfluidic separation module 100 may also separate white bloodcells and CTCs by using various methods such as centrifugation,microfluidic sorting, biochemical methods, etc.). Other methods such asRBC-lysis could also be used to separate red blood cells from the blood.Additional buffers or reagents could also be utilized in the presortingprocess. The resulting presorted contents are then transferred to afluid receiving device 30 to be optically scanned by a scanner 70. Theresulting scanned data 90 is processed by the control unit 80 fordetection and identification of the contents of the biological fluid 16.

Any samples including undesirable constituents 14 may be transferredalong a reprocessing path 62 to be reprocessed or may be isolated alongan isolation path for CTC and CTC-cluster diagnostics (e.g., count,viability, genomic, transcriptomic, molecular, morphological analyses).Any samples which include only desirable constituents 15 may be isolatedseparately and not processed for diagnostics.

N. Example Embodiments

An example embodiment of a biological fluid filtration system 10 maycomprise a receiver path 20 adapted to receive a biological fluid 16from a biological fluid source 17. A fluid receiving device 30 isfluidly connected to the receiver path 20 so as to receive thebiological fluid 16 from the receiver path 20.

A valve 40 may comprise an inlet 41 and a first outlet 42, wherein theinlet 41 of the valve 40 is fluidly connected to the fluid receivingdevice 30. An isolation path 50 may comprise an inlet end, wherein theinlet end of the isolation path 50 is connected to the first outlet 42of the valve 40. A scanner 70 may be oriented toward the fluid receivingdevice 30, with the scanner 70 being adapted to optically scanconstituents 13 of the biological fluid 16 within the fluid receivingdevice 30 so as to derive a scanned data 90 of the constituents 13. Acontrol unit 80 may be communicatively connected to the scanner 70 so asto receive the scanned data 90 from the scanner 70, wherein the controlunit 80 is operatively connected to the valve 40.

The control unit 80 is adapted to compare the scanned data 90 to areference data 91. The reference data 91 may comprise images and/orcharacteristics of desirable constituents 15 such as healthy cells. Thecontrol unit 80 is adapted to switch the valve 40 so as to direct thebiological fluid 16 to the biological fluid source 17 by a return path60 if the scanned data 90 includes only desirable constituents 15meeting the criteria of desirable constituents 15. The control unit 80is adapted to switch the valve 40 so as to direct the biological fluid16 to the isolation path 50 if the scanned data 90 of the biologicalfluid 16 includes one or more undesirable constituents 14 not meetingthe criteria of desirable constituents 15. In another exemplaryembodiment of a biological fluid filtration system 10, the referencedata 91 includes image data representative of desirable constituents 15.

In another exemplary embodiment of a biological fluid filtration system10, the reference data 91 further comprises criteria of optic artifacts,and the control unit 80 is adapted to switch the valve 40 so as todirect the biological fluid 16 to the biological fluid source 17 by thereturn path 60 if the scanned data 90 of the biological fluid 16includes only desirable constituents 15 meeting the criteria of opticartifacts or the criteria of desirable constituents 15.

In another exemplary embodiment of a biological fluid filtration system10, the reference data 91 further comprises recognized criteria ofdesignated pathogens. The control unit 80 is adapted to switch the valve40 so as to direct the biological fluid 16 to the biological fluidsource 17 by the return path 60 if the scanned data 90 of the biologicalfluid 16 includes only desirable constituents 15 meeting the criteria ofdesignated pathogens or the criteria of desirable constituents 15.

The reference data 91 may include image data and/or characteristicsrepresentative of healthy blood constituents. The reference data 91 mayinclude image data and/or characteristics representative oferythrocytes, leukocytes, thrombocytes, and the like.

In another exemplary embodiment, the biological fluid filtration system10, comprises a microfluidic separation module 100, wherein themicrofluidic separation module 100 is adapted to receive blood from thereceiver path 20, generate a primarily leukocyte-rich fluid byseparating leukocytes from other desirable constituents 15 of the blood,and advance the leukocyte-rich fluid with any undesirable components tothe fluid receiving device 30. It should also be appreciated that, inembodiments in which a biological fluid 16 other than blood is beingscanned, the components which are separated during presorting will bydefinition be comprised of different components than with blood. Forexample, in embodiments in which the biological fluid 16 is comprised ofsaliva, urine, cerebrospinal fluid, lymphatic fluids, or leukapheresisextracts, other constituents of the biological fluid 16 may be separatedthan the exemplary components (leukocytes, etc.) of blood. As anotherexample, other components of the biological fluid 16 may be separated insepsis-like use cases.

The one or more undesirable constituents 14 may be comprised ofcirculating tumor cells and pathogens. The fluid receiving device 30 maybe comprised of one or more microfluidic channels 31. The fluidreceiving device 30 may be comprised of a plurality of microwells, suchas a plurality of microwells arranged in a microwell array 32. Thescanner 70 may be adapted to scan each of the plurality of microwells ofthe microwell array 32.

The fluid receiving device 30 may be comprised of a microfluidic dropletgenerator 34. In such an embodiment, a droplet generator 34 may beutilized to encapsulate cells 18 into droplets 39, with each of thedroplets 39 being scanned by the scanner 70. The droplet generator 34may include a dispersed phase channel 35 and one or more continuousphase channels 36 which converge at a juncture 37 in which the cells 18are encapsulated into the droplets 39. The droplets 39 including theencapsulated cells 18 may then be routed through a scanning channel 38in which each such droplet 39 or groups of droplets 39 are scanned bythe scanner 70.

The valve 40 may comprise a plurality of well valves 40 in one-to-onefluid communication with the plurality of microwells of the microwellarray 32. The control unit 80 is operatively connected to the well valve40 of each of the plurality of microwells of the microwell array 32. Insome example embodiments, pipettes could be utilized to extract contentsof a microwell. In other embodiments, multiple wells in an array 32could share a single valve 40.

In another exemplary embodiment of a biological fluid filtration system10, the fluid receiving device 30 is comprised of a microfluidic channel31. In some embodiments, the fluid receiving device 30 is comprised of aplurality of microfluidic channels 31. The plurality of microfluidicchannels 31 may be arranged in parallel or in series. The scanner 70 maybe adapted to scan each of the plurality of microfluidic channels 31.The valve 40 may comprise a plurality of channel valves 40 in one-to-onefluid communication with the plurality of microfluidic channels 31. Thecontrol unit 80 may be operatively connected to the channel valve 40 ofeach of the plurality of microfluidic channels 31. The valve 40 maycomprise a second outlet 43 in fluid communication with the return path60.

The return path 60 may comprise a return channel that is fluidlyconnected to the biological fluid source 17 such that filteredbiological fluid 16 may be returned to the patient 12. In someembodiments, a drug infuser 130 may be fluidly connected to the returnpath 60 so as to introduce various drugs or treatments in the returnpath 60 to treat the patient 12. The drug infuser 130 may comprise areservoir containing a volume of a drug or treatment that is fluidlyconnected to the return path 60. The drug infuser 130 may utilizevalves, pumps, sensors, and the like to control the dosage infusedwithin the return path 60.

In some embodiments, a reprocessing path 62 may be included so as toallow reprocessing of biological fluids 16 multiple times. In such anembodiment, the reprocessing path 62 may be fluidly connected back tothe fluid receiving device 30 such that processed biological fluid 16may be redirected back to the fluid receiving device 30 for additionalprocessing. It should be appreciated that the reprocessing path 62 maybe included in addition to the return path 60, such that certain samplesof biological fluid 16 may be returned to the patient 12, and certainother samples of biological fluid 16 may instead be returned to thefluid receiving device 30 for additional scanning.

In another exemplary embodiment, a method of filtering a biologicalfluid 16 using the biological fluid filtration system 10 comprises thesteps of: directing the biological fluid 16 to the fluid receivingdevice 30; optically scanning the biological fluid 16 within the fluidreceiving device 30 by the scanner 70 to generate the scanned data 90 ofthe biological fluid 16; comparing the scanned data 90 of the biologicalfluid 16 with the reference data 91 by the control unit 80; returningthe biological fluid 16 to the biological fluid source 17 if the scanneddata 90 includes only desirable constituents 15 meeting the criteria ofdesirable constituents 15; and isolating the biological fluid 16 fromthe biological fluid source 17 if the scanned data 90 of the biologicalfluid 16 includes one or more undesirable constituents 14 not meetingthe criteria of desirable constituents 15.

The method may comprise a therapeutic method and the biological fluidsource 17 may comprise an individual patient 12 having a wide range ofconditions, such as but not limited to cancer or infection with variouspathogens. The method may further comprise obtaining a sample ofbiological fluid 16 of the individual patient 12, wherein the referencedata 91 is generated from the sample.

In another exemplary embodiment, the method may further compriseobtaining samples of biological fluid 16 of one or more individualsother than the individual patient 12, wherein the reference data 91 isgenerated from the samples of biological fluid 16 of the one or moreindividuals other than the individual patient 12. The method may furthercomprise the step of performing diagnostics on the undesirableconstituents 14.

In another exemplary embodiment, a method of filtering a biologicalfluid 16 comprises the steps of: receiving the biological fluid 16 froma biological fluid source 17 by a receiver path 20; transferring thebiological fluid 16 from the receiver path 20 to a fluid receivingdevice 30; optically scanning the biological fluid 16 within the fluidreceiving device 30 by a scanner 70 so as to create a scanned data 90 ofthe biological fluid 16; comparing the scanned data 90 of the biologicalfluid 16 with a reference data 91 by a control unit 80, wherein thereference data 91 comprises criteria including images and/ormorphological parameters of desirable constituents 15; returning thebiological fluid 16 to the biological fluid source 17 by a return path60 if the scanned data 90 of the biological fluid 16 includes onlydesirable constituents 15 meeting the criteria of desirable constituents15; and isolating the biological fluid 16 from the biological fluidsource 17 by an isolation path 50 if the scanned data 90 of thebiological fluid 16 includes one or more undesirable constituents 14 notmeeting the criteria of desirable constituents 15. The reference data 91may include data representative of desirable constituents 15.

In another exemplary embodiment of the method of filtering a biologicalfluid 16 using the biological fluid filtration system 10, the referencedata 91 includes criteria of optic artifacts, and further steps may beperformed comprising returning the biological fluid 16 to the biologicalfluid source 17 by a return path 60 if the scanned data 90 of thebiological fluid 16 includes only desirable constituents 15 meeting thecriteria of optic artifacts or the criteria of desirable constituents15, and isolating the biological fluid 16 from the biological fluidsource 17 by an isolation path 50 if the scanned data 90 of thebiological fluid 16 includes one or more undesirable constituents 14 notmeeting the criteria of optic artifacts or the criteria of desirableconstituents 15.

In another exemplary embodiment of the method of filtering a biologicalfluid 16 using the biological fluid filtration system 10, the referencedata 91 includes criteria including images and/or morphologicalparametric data representative of designated pathogens, and furthersteps may be performed comprising returning the biological fluid 16 tothe biological fluid source 17 by a return path 60 if the scanned data90 of the biological fluid 16 includes only desirable constituents 15meeting the criteria of optic artifacts/pathogens or the criteria ofdesirable constituents 15, and isolating the biological fluid 16 fromthe biological fluid source 17 by an isolation path 50 if the scanneddata 90 of the biological fluid 16 includes one or more undesirableconstituents 14 not meeting the criteria of optic artifacts/pathogens orthe criteria of desirable constituents 15.

The reference data 91 may include image data or morphologicalcharacteristics representative of healthy blood constituents. By way ofexample and without limitation, the reference data 91 may include datarepresentative of erythrocytes, leukocytes, or thrombocytes.

In another exemplary embodiment of the method of filtering a biologicalfluid 16 using the biological fluid filtration system 10, the method mayfurther comprise the steps of: sorting the biological fluid 16 from thereceiver path 20 to separate leukocytes from other desirableconstituents 15 of the biological fluid 16 to generate a leukocyte-richfluid, and transferring the leukocyte-rich fluid to the fluid receivingdevice 30. The one or more undesirable constituents 14 may be comprisedof circulating tumor cells, clusters thereof, or various otherpathogens.

In another exemplary embodiment of the method of filtering a biologicalfluid 16 using the biological fluid filtration system 10, each of theplurality of microwells of the microwell array 32 may comprise a wellvalve 40 having a first port fluidly connected to the isolation path 50and a second port fluidly connected to the return path 60. In otherembodiments, each of the plurality of microwells of the microwell array32 is fluidly connected via a well valve 40 having a single port, withthe port being fluidly and selectively connected to both the return path60 and the isolation path 50.

The control unit 80 may be operatively connected to the well valve 40 ofeach of the plurality of microwells of the microwell array 32. The fluidreceiving device 30 may be comprised of a microfluidic channel 31 or aplurality of microfluidic channels 31. The plurality of microfluidicchannels 31 may be arranged in a parallel series. The scanner 70 may beadapted to scan each of the plurality of microfluidic channels 31. Insome example embodiments, pipettes could be utilized to extract contentsof a microwell. In other embodiments, multiple wells in an array couldshare a single valve 40.

In another exemplary embodiment of the method of filtering a biologicalfluid 16 using the biological fluid filtration system 10, each of theplurality of microfluidic channels 31 may comprise a channel valve 40having a first port fluidly connected to the isolation path 50 and asecond port fluidly connected to the return path 60. The control unit 80may be operatively connected to the channel valve 40 of each of theplurality of microfluidic channels 31.

In another exemplary embodiment, a method of filtering a biologicalfluid 16 comprises the steps of: receiving the biological fluid 16 froma biological fluid source 17; separating the biological fluid 16 into afirst portion, a second portion, and a third portion, the first portioncomprising only desirable constituents 15, the second portion comprisinga mix of undesirable and desirable constituents 14, 15, and the thirdportion comprising only undesirable constituents 14; returning the firstportion of the biological fluid 16 to the biological fluid source 17 bya return path 60; isolating the third portion of the biological fluid 16from the biological fluid source 17 by an isolation path 50;transferring the second portion of the biological fluid 16 to a fluidreceiving device 30; optically scanning the second portion of thebiological fluid 16 within the fluid receiving device 30 by a scanner 70so as to create a scanned data 90 of the second portion of thebiological fluid 16; comparing the scanned data 90 of the second portionof the biological fluid 16 with a reference data 91 by a control unit80, the reference data 91 comprising criteria of desirable constituents15; returning the second portion of the biological fluid 16 to thebiological fluid source 17 by the return path 60 if the scanned data 90of the biological fluid 16 includes only desirable constituents 15meeting the criteria of desirable constituents 15; and isolating thesecond portion of the biological fluid 16 from the biological fluidsource 17 by the isolation path 50 if the scanned data 90 of thebiological fluid 16 includes one or more constituents 13 not meeting thecriteria of desirable constituents 15. The step of separating thebiological fluid 16 may be comprised of dielectric sorting, Dean flowfractionation, or hemodynamic properties.

In another exemplary embodiment of the method of filtering a biologicalfluid 16 using the biological fluid filtration system 10, multiplescanners 70 may be used simultaneously to optically scan the fluidreceiving device 30. In such an embodiment, different areas, such asdifferent microwells of a microwell array 32, different microfluidicchannels 31, or different droplets 39 of a droplet generator 34, may beoptically scanned by a plurality of scanners 70 rather than a singularscanner 70.

In another exemplary embodiment of the method of filtering a biologicalfluid 16 using the biological fluid filtration system 10, the fluidreceiving device 30 may be scanned multiple times by the scanner 70before transfer to the return path 60. Such an embodiment will utilizemultiple scans to ensure and verify that the biological fluid 16 beingscanned does not include any undesirable constituents 14 prior to bereleased to the return path 60. Such an embodiment will reduce the risksof inaccurate scans by verifying through multiple scans prior toreleasing the biological fluid 16.

Unless otherwise defined, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this invention belongs. Although methods and materialssimilar to or equivalent to those described herein can be used in thepractice or testing of the biological fluid filtration system, suitablemethods and materials are described above. All patent applications,patents, and printed publications cited herein are incorporated hereinby reference in their entireties, except for any definitions, subjectmatter disclaimers or disavowals, and except to the extent that theincorporated material is inconsistent with the express disclosureherein, in which case the language in this disclosure controls. Thebiological fluid filtration system may be embodied in other specificforms without departing from the spirit or essential attributes thereof,and it is therefore desired that the present embodiment be considered inall respects as illustrative and not restrictive. Any headings utilizedwithin the description are for convenience only and have no legal orlimiting effect.

What is claimed is:
 1. A portable biological fluid filtration system,comprising: a housing including an inlet and an outlet, wherein theinlet of the housing is adapted to receive a biological fluid from apatient; a fluid filtration device including an inlet, a first outlet,and a second outlet, wherein the inlet of the fluid filtration device isfluidly connected to the inlet of the housing, wherein the fluidfiltration device is adapted to receive the biological fluid, andwherein the second outlet of the fluid filtration device is fluidlyconnected to the outlet of the housing; wherein the fluid filtrationdevice is configured to separate a first subset of constituents and asecond subset of constituents from the biological fluid, wherein thefirst subset of constituents includes an undesirable constituent,wherein the second subset of constituents does not include theundesirable constituent, wherein the fluid filtration device isconfigured to direct the first subset of constituents through the firstoutlet of the fluid filtration device, and wherein the fluid filtrationdevice is configured to direct the second subset of constituents throughthe second outlet of the fluid filtration device; and a cartridgeremovably attached to the housing, wherein the cartridge is fluidlyconnected to the first outlet of the fluid filtration device, whereinthe cartridge is adapted to receive the first subset of constituents. 2.The portable biological fluid filtration system of claim 1, furthercomprising a return channel fluidly connected between the outlet of thehousing and a source of the biological fluid.
 3. The portable biologicalfluid filtration system of claim 1, wherein the fluid filtration deviceincludes a valve, wherein the valve is comprised of an inlet, a firstoutlet, and a second outlet, wherein the inlet of the valve is fluidlyconnected to the inlet of the fluid filtration device, wherein the firstoutlet of the valve is fluidly connected to the first outlet of thefluid filtration device, and wherein the second outlet of the valve isfluidly connected to the second outlet of the fluid filtration device.4. The portable biological fluid filtration system of claim 3, whereinthe fluid filtration device includes a scanner configured to scan thebiological fluid within the fluid filtration device to produce a scanneddata relating to the biological fluid within the fluid filtrationdevice.
 5. The portable biological fluid filtration system of claim 4,wherein the fluid filtration device includes a control unit incommunication with the scanner and the valve, wherein the control unitis configured to receive the scanned data from the scanner, and whereinthe control unit is configured to control the valve based on the scanneddata from the scanner.
 6. The portable biological fluid filtrationsystem of claim 1, wherein the fluid filtration device includes ananti-coagulant insert, wherein the anti-coagulant insert is configuredto apply an anti-coagulant to the biological fluid within the fluidfiltration device.
 7. The portable biological fluid filtration system ofclaim 1, wherein the fluid filtration device includes a power source. 8.The portable biological fluid filtration system of claim 1, wherein thehousing includes at least one indicator adapted to convey an operationalstatus of the fluid filtration device.
 9. The portable biological fluidfiltration system of claim 1, wherein the fluid filtration deviceincludes a buffer fluid insert, wherein the buffer fluid insert isadapted to apply a buffer fluid to the biological fluid within the fluidfiltration device.
 10. The portable biological fluid filtration systemof claim 1, wherein the housing includes a presorting device fluidlyconnected between the inlet of the housing and the inlet of the fluidfiltration device.
 11. The portable biological fluid filtration systemof claim 10, wherein the presorting device is comprised of a centrifuge.12. The portable biological fluid filtration system of claim 10, whereinthe presorting device is adapted to apply a chemical agent to thebiological fluid.
 13. The portable biological fluid filtration system ofclaim 12, wherein the chemical agent is comprised of a red blood celllysis buffer.
 14. The portable biological fluid filtration system ofclaim 10, wherein the presorting device is comprised of a microfluidicseparation device.
 15. The portable biological fluid filtration systemof claim 14, wherein the microfluidic separation device is adapted toapply inertial focusing to the biological fluid.
 16. The portablebiological fluid filtration system of claim 1, further comprising a druginfuser device fluidly connected between the second outlet of the fluidfiltration device and the outlet of the housing.
 17. The portablebiological fluid filtration system of claim 1, further comprising acatheter including an inlet port and an outlet port, wherein the inletport of the catheter is fluidly connected to the inlet of the housing,and wherein the outlet port of the catheter is fluidly connected to theoutlet of the housing.
 18. A method of filtering a biological fluidusing the portable biological fluid filtration system of claim 1,comprising the steps of: receiving the biological fluid by the fluidfiltration device; separating the first subset of constituents from thesecond subset of constituents by the fluid filtration device; directingthe first subset of constituents into the cartridge; and directing thesecond subset of constituents through the outlet of the housing.
 19. Theportable biological fluid filtration system of claim 1, wherein thehousing is adapted to be worn on a body of a patient.
 20. A portablebiological fluid filtration system, comprising: a housing including aninlet and an outlet, wherein the inlet of the housing is adapted toreceive a biological fluid from a patient, wherein the housing isadapted to be worn on a body of a patient; a fluid filtration deviceincluding an inlet, a first outlet, and a second outlet, wherein theinlet of the fluid filtration device is fluidly connected to the inletof the housing, wherein the fluid filtration device is adapted toreceive the biological fluid, and wherein the second outlet of the fluidfiltration device is fluidly connected to the outlet of the housing;wherein the fluid filtration device is configured to separate a firstsubset of constituents and a second subset of constituents from thebiological fluid, wherein the first subset of constituents includes anundesirable constituent, wherein the second subset of constituents doesnot include the undesirable constituent, wherein the fluid filtrationdevice is configured to direct the first subset of constituents throughthe first outlet of the fluid filtration device, and wherein the fluidfiltration device is configured to direct the second subset ofconstituents through the second outlet of the fluid filtration device;wherein the fluid filtration device includes a valve, wherein the valveis comprised of an inlet, a first outlet, and a second outlet, whereinthe inlet of the valve is fluidly connected to the inlet of the fluidfiltration device, wherein the first outlet of the valve is fluidlyconnected to the first outlet of the fluid filtration device, andwherein the second outlet of the valve is fluidly connected to thesecond outlet of the fluid filtration device; wherein the fluidfiltration device includes a scanner configured to scan the biologicalfluid within the fluid filtration device to produce a scanned datarelating to the biological fluid within the fluid filtration device;wherein the fluid filtration device includes a control unit incommunication with the scanner and the valve, wherein the control unitis configured to receive the scanned data from the scanner, and whereinthe control unit is configured to control the valve based on the scanneddata from the scanner; and a cartridge removably attached to thehousing, wherein the cartridge is fluidly connected to the first outletof the fluid filtration device, wherein the cartridge is adapted toreceive the first subset of constituents.